WO2004063029A2 - Droplet dispensation from a reservoir with reduction in uncontrolled electrostatic charge - Google Patents

Droplet dispensation from a reservoir with reduction in uncontrolled electrostatic charge Download PDF

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Publication number
WO2004063029A2
WO2004063029A2 PCT/US2004/000522 US2004000522W WO2004063029A2 WO 2004063029 A2 WO2004063029 A2 WO 2004063029A2 US 2004000522 W US2004000522 W US 2004000522W WO 2004063029 A2 WO2004063029 A2 WO 2004063029A2
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WO
WIPO (PCT)
Prior art keywords
reservoir
fluid
droplet
electrostatic charge
acoustic
Prior art date
Application number
PCT/US2004/000522
Other languages
French (fr)
Other versions
WO2004063029A3 (en
Inventor
Mitchell W. Mutz
David Soong-Hua Lee
George Mclendon
Original Assignee
Picoliter Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US10/340,557 external-priority patent/US7070260B2/en
Priority claimed from US10/668,534 external-priority patent/US6916083B2/en
Application filed by Picoliter Inc. filed Critical Picoliter Inc.
Priority to AT04701230T priority Critical patent/ATE554931T1/en
Priority to EP04701230A priority patent/EP1585636B1/en
Publication of WO2004063029A2 publication Critical patent/WO2004063029A2/en
Publication of WO2004063029A3 publication Critical patent/WO2004063029A3/en

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Classifications

    • BPERFORMING OPERATIONS; TRANSPORTING
    • B41PRINTING; LINING MACHINES; TYPEWRITERS; STAMPS
    • B41JTYPEWRITERS; SELECTIVE PRINTING MECHANISMS, i.e. MECHANISMS PRINTING OTHERWISE THAN FROM A FORME; CORRECTION OF TYPOGRAPHICAL ERRORS
    • B41J2/00Typewriters or selective printing mechanisms characterised by the printing or marking process for which they are designed
    • B41J2/005Typewriters or selective printing mechanisms characterised by the printing or marking process for which they are designed characterised by bringing liquid or particles selectively into contact with a printing material
    • B41J2/01Ink jet
    • B41J2/135Nozzles
    • B41J2/14Structure thereof only for on-demand ink jet heads
    • B41J2/14008Structure of acoustic ink jet print heads

Definitions

  • This invention relates generally to devices and methods for accurately dispensing a droplet from a reservoir, optionally toward a substrate, wherein the volume and/or trajectory of the droplet do not substantially deviate from a predetermined volume and/or trajectory. More particularly, the invention relates to devices and methods for reducing the uncontrolled electrostatic charges that can alter the volume and/or trajectory of a droplet, which is typically ejected through the application of focused acoustic radiation.
  • U.S. Patent No. 4,308,547 to Lovelady et al. describes a liquid drop emitter that utilizes acoustic principles to eject droplets from a body of liquid onto a moving document to result in the formation of characters or barcodes thereon.
  • a nozzleless inkjet printing apparatus is used such that controlled drops of ink are propelled by an acoustical force produced by a curved transducer at or below the surface of the ink.
  • the device includes an acoustic radiation generator for generating acoustic and a focusing means, e.g., a curved surface, for focusing acoustic radiation generated by the generator.
  • a focusing means e.g., a curved surface
  • the acoustic generator is acoustically coupled to the reservoir and activated to generate acoustic radiation.
  • the focusing means then focuses the generated acoustic radiation at a point near a free fluid surface within the fluid contained in the reservoir. As a result, a fluid droplet is ejected from reservoir.
  • U.S. Patent No. 6,596,239 to Williams describes technologies that employ focused acoustic technology as well.
  • Acoustic ejection provides a number of advantages over other fluid dispensing technologies.
  • nozzleless fluid ejection devices are not subject to clogging and their associated disadvantages, e.g., misdirected fluid or improperly sized droplets.
  • focused acoustic radiation may be used to effect nozzleless fluid ejection, and devices using focused acoustic radiation are not generally subject to clogging and the disadvantages associated therewith, e.g., misdirected fluid or improperly sized droplets.
  • acoustic technology does not require the use of capillaries or involve invasive mechanical actions, for example, those associated with the introduction of a pipette tip into a reservoir of fluid.
  • Acoustic radiation may also be used to assess the contents of one or more reservoirs.
  • the device described in U.S. Patent No. 6,666,541 to Ellson et al. may also be used to produce a detection acoustic wave that is transmitted to the fluid surface of the reservoir to become a reflected acoustic wave. Characteristics of the reflected acoustic radiation may then be analyzed in order to assess the spatial relationship between the acoustic radiation generator and the fluid surface.
  • pool depth feedback technology using acoustic radiation is described in U.S. Patent No. 5,520,715 to Oeftering.
  • U.S. Patent No. 6,596,239 to Williams describes acoustic ejection and pool depth detection technology.
  • a generator for generating acoustic radiation is placed in acoustic coupling relationship with the reservoir.
  • the generator may be placed within the reservoir to establish acoustic coupling, e.g., submerged in a fluid contained in the reservoir, submersion is undesirable when the acoustic generator is used to eject different fluids in rapid succession. Cleaning would be required to avoid contamination between the fluids.
  • a preferred approach is to couple the generator to an exterior surface of the reservoir and to avoid placing the generator in the reservoir. As a result, the generator does not contact any fluid that the reservoir may contain.
  • acoustic coupling may be achieved between an acoustic generator and a reservoir via an acoustic coupling medium.
  • a coupling medium allows transmission of acoustic radiation therethrough and into the reservoir.
  • the acoustic coupling medium is an acoustically homogeneous fluid in conformal contact with both acoustic generator and the reservoir.
  • fluids used in pharmaceutical, biotechnological, and other scientific industries may be rare and/or expensive, techniques capable of handling small volumes of fluids provide readily apparent advantages over those requiring relatively larger volumes.
  • fluids for use in combinatorial methods are provided as a collection or library of organic and or biological compounds.
  • well plates are used to store a large number of fluids for screening and/or processing.
  • Well plates are typically of single piece construction and comprise a plurality of identical wells, wherein each well is adapted to contain a small volume of fluid.
  • Such well plates are commercially available in standardized sizes and may contain, for example, 96, 384, 1536, or 3456 wells per well plate.
  • the ideal fluid-dispensing technique for pharmaceutical, biotechnological, medical (including clinical testing), and other industries provides for highly repeatable and accurate ejection of minute volumes of fluids directly from wells of a well plate.
  • the dispensing technique provides for deposition of droplets on a substrate surface, wherein droplet volume—and thus "spot" size on the substrate surface- can be carefully controlled.
  • the droplets In order to ensure accurate placement of the droplets on a substrate surface, the droplets must take an appropriate trajectory from the wells of well plates towards the destination substrate.
  • U.S. Patent Nos. 6,079,814 and 6,367,909 each to Lean et al., describe printing methods and apparatuses that employ electric fields to reduce drop placement errors.
  • an aperture plate is used to charge a free surface of a fluid in a reservoir. Then, focused acoustic radiation is applied to a point near the fluid surface so as to eject a charged droplet therefrom and through the aperture of the plate. Additional electric fields may be employed to direct the charged droplet so that it follows a predetermined trajectory.
  • an electric field may also serve to tack a recording medium in position to receive the ink droplet.
  • the invention provides a device comprised of a reservoir adapted to contain a fluid and a dispenser for dispensing a fluid droplet from the reservoir.
  • a means is employed for reducing uncontrolled electrostatic charge on the reservoir when the reservoir is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom.
  • the means for reducing uncontrolled electrostatic charge is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory. Often grounding is used to reduce or eliminate uncontrolled electrostatic charge.
  • the invention provides a similar device that further comprises a substrate positioned to receive the dispensed droplet.
  • a substrate positioned to receive the dispensed droplet.
  • a means for reducing uncontrolled electrostatic charge is provided that is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
  • the dispenser is comprised of an acoustic ejector.
  • the acoustic ejector may comprise an acoustic radiation generator for generating acoustic radiation and a focusing means for focusing the acoustic radiation generated.
  • the invention also provides a means for positioning the ejector in acoustic coupling relationship to the reservoir.
  • the reservoir, the substrate, and any other component of the device prone to accumulate uncontrolled electrostatic charge have an electrical resistivity of no more than about 10 11 ohm-cm, have a surface electrical resistivity of no more than about 10 12 ohm/sq, or both. This may be achieved by using a material that is at least partially nonmetallic or polymeric.
  • the invention provides a method for dispensing a droplet from a reservoir containing a fluid.
  • the method involves reducing uncontrolled electrostatic charge on the reservoir when the reservoir is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom.
  • uncontrolled electrostatic charge is reduced to a level effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
  • the invention provides a method for dispensing a droplet from a reservoir containing a fluid onto a substrate.
  • the method involves reducing uncontrolled electrostatic charge on the reservoir and/or the substrate when the reservoir and/or substrate are prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of the dispensed droplet.
  • Uncontrolled electrostatic charge is reduced to a level effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
  • focused acoustic radiation may be applied in a manner effective to eject a droplet of fluid from the reservoir.
  • FIG. 1A and FIG. IB collectively referred to as FIG. 1, schematically illustrate in simplified cross-sectional view the operation of a focused acoustic ejection device in the preparation of a plurality of features on a substrate surface.
  • FIG. 1A shows the acoustic ejector acoustically coupled to a first reservoir and having been activated in order to eject a first droplet of fluid from within the reservoir toward a particular site on a substrate surface.
  • FIG. IB shows the acoustic ejector acoustically coupled to a second reservoir and having been activated to eject a second droplet of fluid from within the second reservoir.
  • FIG. 2 illustrates in cross-sectional schematic view the ejection of droplets of fluid from a volume of fluid on a substrate surface into an inlet opening disposed on a terminus of a capillary.
  • FIG. 3A, FIG. 3B, FIG. 3C and FIG. 3D collectively referred to as FIG. 3, schematically illustrate in simplified cross-sectional view a known device that employs an acoustic coupling fluid and the disadvantages associated therewith.
  • the device comprises first and second reservoirs, a combined acoustic analyzer and ejector unit, and an ejector positioning means.
  • FIG. 3A shows the acoustic unit acoustically coupled to the first reservoir so that the unit is activated to determine the position of the free fluid surface within the first reservoir.
  • FIG. 3A shows the acoustic unit acoustically coupled to the first reservoir so that the unit is activated to determine the position of the free fluid surface within the first reservoir.
  • FIG. 3B depicts the repositioning of the acoustic unit toward the reservoir and the activation acoustic unit in order to eject a droplet of fluid from within the first reservoir toward a site on a substrate surface to form an array.
  • FIG. 3C shows the acoustic unit acoustically coupled to the second reservoir so that the unit is activated to determine the position of the free fluid surface within the second reservoir.
  • FIG. 3D depicts the repositioning of the acoustic unit away from the reservoir and the activation acoustic unit in order to eject a droplet of fluid from within the second reservoir toward a site on a substrate surface.
  • FIG. 4 schematically illustrate in simplified cross-sectional view a device that includes a nozzle located within a collector.
  • FIG. 5 schematically illustrates in simplified cross-sectional view an acoustic device having a dispenser that employs a stationary opposing piston design.
  • FIG. 6 schematically illustrates in simplified cross-sectional view an acoustic device similar to that of FIG. 3 except that the acoustic ejector and the positioning means are sealed and in a container filled completely with the acoustic coupling fluid
  • acoustic coupling and "acoustically coupled” as used herein refer to a state wherein an object is placed in direct or indirect contact with another object so as to allow acoustic radiation to be transferred between the objects without substantial loss of acoustic energy.
  • an "acoustic coupling medium” is needed to provide an intermediary through which acoustic radiation may be transmitted.
  • an ejector may be acoustically coupled to a fluid, e.g., by immersing the ejector in the fluid or by interposing an acoustic coupling medium between the ejector and the fluid, in order to transfer acoustic radiation generated by the ejector through the acoustic coupling medium and into the fluid.
  • the acoustic coupling medium may include or be formed entirely of a solid material.
  • array refers to a two-dimensional arrangement of features, such as an arrangement of reservoirs (e.g., wells in a well plate) or an arrangement of different moieties, including ionic, metallic, or covalent crystalline, e.g., molecular crystalline, composite, ceramic, vitreous, amorphous, fluidic, or molecular materials on a substrate surface (as in an ohgonucleotide or peptidic array).
  • Arrays are generally comprised of regular, ordered features, as in, for example, a rectilinear grid, parallel stripes, spirals, and the like, but non-ordered arrays may be advantageously used as well.
  • rectilinear array refers to an array that has rows and columns of features wherein the rows and columns typically, but not necessarily, intersect each other at a ninety- degree angle.
  • An array is distinguished from the more general term "pattern” in that patterns do not necessarily contain regular and ordered features. Arrays typically but do not necessarily comprise at least about 4 to about 10,000,000 features, generally in the range of about 4 to about 1,000,000 features.
  • the term "attenuation” is used herein in its ordinary sense and refers to the decrease in intensity of a wave due to scattering and/or absorption of energy. Typically, attenuation occurs with little or no distortion but does not include intensity reduction due to geometric spreading. Thus, the term “attenuation coefficient” refers to the rate of diminution of wave intensity with respect to distance along a transmission path.
  • biomolecule and “biological molecule” are used interchangeably herein to refer to any organic molecule that is, was, or can be a part of a living organism, regardless of whether the molecule is naturally occurring, recombinantly produced, or chemically synthesized in whole or in part.
  • the terms encompass, for example, nucleotides, amino acids, and monosaccharides, as well as oligomeric and polymeric species, such as oligonucleotides and polynucleotides; peptidic molecules, such as oligopeptides, polypeptides, and proteins; saccharides, such as disaccharides, oligosaccharides, polysaccharides, and mucopolysaccharides or peptidoglycans (peptido-polysaccharides); and the like.
  • the terms also encompass ribosomes, enzyme cofactors, pharmacologically active agents, and the like. Additional information relating to the term "biomolecule" can be found in U.S. Patent No. 6,666,541 to Ellson et al.
  • capillary is used herein to refer to a conduit having a bore of small dimension.
  • capillaries for electrophoresis that are free standing tubes have an inner diameter in the range of about 50 to about 250 ⁇ m.
  • Capillaries with extremely small bores integrated to other devices, such as openings for loading microchannels of microfluidic devices can be as small as 1 ⁇ m, but in general these capillary openings are in the range of about 10 to about 100 ⁇ m.
  • the inner diameter of capillaries may range from about 0.1 to about 3 mm and preferably from about 0.5 to about 1 mm.
  • a capillary can represent a portion of a microfluidic device.
  • the capillary may be an integral or affixed (permanently or detachably) portion of the microfluidic device.
  • fluid refers to matter that is nonsolid, or at least partially gaseous and/or liquid, but not entirely gaseous.
  • a fluid may contain a solid that is minimally, partially, or fully solvated, dispersed, or suspended.
  • fluids include, without limitation, aqueous liquids (including waterier se and salt water) and nonaqueous liquids such as organic solvents and the like.
  • aqueous liquids including waterier se and salt water
  • nonaqueous liquids such as organic solvents and the like.
  • the term “fluid” is not synonymous with the term "ink” in that an ink must contain a colorant and may not be gaseous.
  • focusing means and "acoustic focusing means” refer to a means for causing acoustic waves to converge at a focal point, either by a device separate from the acoustic energy source that acts like an optical lens, or by the spatial arrangement of acoustic energy sources to effect convergence of acoustic energy at a focal point by constructive and destructive interference.
  • a focusing means may be as simple as a solid member having a curved surface, or it may include complex structures such as those found in Fresnel lenses, which employ diffraction in order to direct acoustic radiation. Suitable focusing means also include phased array methods as are known in the art and described, for example, in U.S. Patent No.
  • library and “combinatorial library” are used interchangeably herein to refer to a plurality of chemical or biological moieties arranged in a pattern or an array such that the moieties are individually addressable.
  • the plurality of chemical or biological moieties is present on the surface of a substrate, and in other instances the plurality of moieties represents the contents of a plurality of reservoirs.
  • each moiety is different from each of the other moieties.
  • the moieties may be, for example, peptidic molecules and/or oligonucleotides.
  • the "limiting dimension" of an opening refers herein to the theoretical maximum diameter of a sphere that can pass through an opening without deformation.
  • the limiting dimension of a circular opening is the diameter of the opening.
  • the limiting dimension of a rectangular opening is the length of the shorter side of the rectangular opening.
  • the opening may be present on any solid body including, but not limited to, sample vessels, substrates, capillaries, microfluidic devices, and ionization chambers. Depending on the purpose of the opening, the opening may represent an inlet and/or an outlet.
  • molecular fragment refers to any particular composition of matter, e.g., a molecular fragment, an intact molecule (including a monomeric molecule, an oligomeric molecule, or a polymer), or a mixture of materials (for example, an alloy or a laminate).
  • the term "near,” as used herein, refers to the distance from the focal point of the focused acoustic radiation to the surface of the fluid from which a droplet is to be ejected, and indicates that the distance should be such that the focused acoustic radiation directed into the fluid results in droplet ejection from the fluid surface; one of ordinary skill in the art will be able to select an appropriate distance for any given fluid using straightforward and routine experimentation.
  • a suitable distance between the focal point of the acoustic radiation and the fluid surface is in the range of about 1 to about 15 times the wavelength of the acoustic radiation in the fluid, more typically in the range of about 1 to about 10 times that wavelength, preferably in the range of about 1 to about 5 times that wavelength.
  • radiation is used in its ordinary sense and refers to emission and propagation of energy in the form of a waveform disturbance traveling through a medium such that energy is transferred from one particle of the medium to another, generally without causing any permanent displacement of the medium itself.
  • radiation may refer, for example, to electromagnetic waveforms as well as acoustic vibrations.
  • acoustic radiation and “acoustic energy” are used interchangeably herein and refer to the emission and propagation of energy in the form of sound waves.
  • acoustic radiation may be focused using a focusing means, as discussed below.
  • acoustic radiation may have a single frequency and associated wavelength
  • acoustic radiation may take a form, e.g. a "linear chirp,” that includes a plurality of frequencies.
  • characteristic wavelength is used to describe the mean wavelength of acoustic radiation having a plurality of frequencies.
  • a fluid contained in a reservoir necessarily will have a free surface, e.g., a surface that allows acoustic radiation to be reflected therefrom or a surface from which a droplet may be acoustically ejected.
  • a reservoir may also be a locus on a substrate surface within which a fluid is constrained.
  • substrate refers to any material having a surface onto which one or more fluids may be deposited.
  • the substrate may be constructed in any of a number of forms including, for example, wafers, slides, well plates, or membranes.
  • the substrate may be porous or nonporous as required for deposition of a particular fluid.
  • Suitable substrate materials include, but are not limited to, supports that are typically used for solid phase chemical synthesis, such as polymeric materials (e.g., polystyrene, polyvinyl acetate, polyvinyl chloride, polyvinyl pyrrolidone, polyacrylonitrile, polyacrylamide, polymethyl methacrylate, polytetrafluoroethylene, polyethylene, polypropylene, polyvinylidene fluoride, polycarbonate, and divinylbenzene styrene-based polymers), agarose (e.g., Sepharose®), dextran (e.g., Sephadex®), cellulosic polymers and other polysaccharides, silica and silica-based materials, glass (particularly controlled pore glass, or "CPG") and functionalized glasses, ceramics, and such substrates treated with surface coatings, e.g., with microporous polymers (particularly cellulosic polymers such as nitrocellulose), microporous metallic compounds (
  • substantially refers to a volume that does not deviate by more than about 25%, preferably 10%, more preferably 5%, and most preferably at most 2%, from the predetermined volume. Other uses of the term “substantially” involve an analogous definition.
  • sample vessel refers to any hollow or concave receptacle having a structure that allows for sample processing and/or analysis.
  • a sample vessel has an inlet opening through which sample may be introduced and an optional, but preferred, outlet opening through which processed or analyzed sample may exit.
  • the invention relates to devices and methods for dispensing a fluid droplet of a predetermined volume and/or predetermined trajectory from a reservoir adapted to contain a fluid.
  • the invention derives from the observation that fluid dispensing devices or components thereof sometimes accumulate uncontrolled electrostatic charge such that droplets dispensed therefrom exhibit a volume and/or trajectory that substantially deviate from the predetermined volume and/or predetermined trajectory. This is particularly problematic when the device is adapted to dispense droplets containing a minute volume of fluid. Often, the reservoir itself is prone to accumulate such uncontrolled electrostatic charge.
  • the invention provides for the reduction of such uncontrolled electrostatic charge in a manner effective to ensure that the volume and/or trajectory of the dispensed droplet conform to the predetermined volume and/or trajectory.
  • the invention is particularly suited for applications that require the efficient transport and/or deposition of small quantities of fluid.
  • the triboelectric effect by which an item will typically accumulate uncontrolled electrostatic charge through friction, pressure, and separation.
  • the magnitude of the static charge is typically determined by material composition, applied forces, separation rate, and dissipative forces.
  • the ability of a material to surrender or gain electrons is a function of the conductivity of the material.
  • the tendency of a material to accumulate uncontrolled electrostatic charge is inversely correlated to the surface and/or volume conductivity of the material. Accordingly, the invention is particularly suited for use in devices comprised of components that exhibit a low electrical conductivity or high electrical resistivity.
  • the invention will be useful to reduce uncontrolled electrostatic charge in items having a volume electrical resistivity of at least 10 ohm-cm and/or a surface electrical resistivity of at least 10 14 ohm/sq.
  • a volume electrical resistivity of at least 10 15 or 10 16 ohm-cm and/or a surface electrical resistivity of at least 10 16 or 10 17 ohm/sq will be particularly useful to discharge items having a volume electrical resistivity of at least 10 15 or 10 16 ohm-cm and/or a surface electrical resistivity of at least 10 16 or 10 17 ohm/sq.
  • the invention may be employed with any type of fluid dispenser that serves to dispense one or more droplets of fluid from a reservoir. Any fluid droplet dispensing techniques known in the art may be used in conjunction with the present invention.
  • the invention may be used with dispensers such as inkjet printheads (both thermal and piezoelectric), pipettes, capillaries, syringes, displacement pumps, rotary pumps, peristaltic pumps, vacuum devices, flexible or rigid tubing, valves, manifolds, pressurized gas canisters, and combinations thereof.
  • an ejector may be acoustically coupled to a reservoir containing a fluid in order to eject a droplet therefrom.
  • the reservoir may be a well of a well plate.
  • this device configuration allows droplets to be ejected from near the base of a well, uncontrolled electrostatic charge anywhere in the well, e.g., the base or sidewalls, may have a strong effect influence on the volume and or trajectory of such droplets. Since current inkjet systems do not typically exhibit such a configuration, devices having such a configuration, e.g., devices that employ focused acoustic radiation, may benefit more from the invention than ordinary inkjet technologies.
  • one embodiment of the invention provides a device for acoustically ejecting a droplet of fluid from a reservoir.
  • the device is comprised of a reservoir adapted to contain a fluid, an ejector for ejecting a droplet from the reservoir, and a means for positioning the ejector in acoustic coupling relationship to the reservoir.
  • the ejector comprises an acoustic radiation generator for generating acoustic radiation and a focusing means for focusing the acoustic radiation generated by the generator.
  • the acoustic radiation is focused at a focal point within and sufficiently near the fluid surface in the reservoir to result in the ejection of droplets therefrom. Furthermore, a means is provided for reducing any uncontrolled electrostatic charge on the device or a portion thereof that alters the volume and or trajectory of a droplet ejected from the reservoir. As a result, the volume and/or trajectory of the ejected droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
  • the device may be constructed to include the reservoir as an integrated or permanently attached component of the device. However, to provide modularity and interchangeability of components, it is preferred that the device be constructed with a removable reservoir. Optionally, a plurality of reservoirs many be provided. Generally, the reservoirs are arranged in a pattern or an array to provide each reservoir with individual systematic addressability. In addition, while each of the reservoirs may be provided as a discrete or stand-alone item, in circumstances that require a large number of reservoirs, it is preferred that the reservoirs be attached to each other or represent integrated portions of a single reservoir unit. For example, the reservoirs may represent individual wells in a well plate.
  • Many well plates suitable for use with the device are commercially available and may contain, for example, 96, 384, 1536, or 3456 wells per well plate, having a full skirt, half skirt, or no skirt.
  • the wells of such well plates typically form rectilinear arrays.
  • Manufacturers of suitable well plates for use in the employed device include Corning, Inc. (Corning, New York) and Greiner America, Inc. (Lake Mary, Florida).
  • the availability of such commercially available well plates does not preclude the manufacture and use of custom-made well plates containing at least about 10,000 wells, or as many as 100,000 to 500,000 wells, or more.
  • the wells of such custom-made well plates may form rectilinear or other types of arrays.
  • the Society for Biomolecular Screening (Danbury, Connecticut) has formed the Microplate Standards Development Committee to recommend and maintain standards to facilitate the automated processing of small volume well plates on behalf of and for acceptance by the American National Standards Institute.
  • reservoirs or wells must be compatible with DMSO.
  • a number of materials are suitable for the construction of reservoirs and include, but are not limited to, ceramics such as silicon oxide and aluminum oxide, metals such as stainless steel and platinum, and polymers such as polyester and polytetrafluoroethylene.
  • the reservoirs may be constructed from an optically opaque material that has sufficient acoustic transparency for substantially unimpaired functioning of the device.
  • the reservoir may be adapted to contain any type of fluid, metallic or nonmetallic, organic or inorganic.
  • polymeric materials are particularly suited for use in forming reservoirs for use with the invention, e.g., well plates that conform to industrial standards. Such materials typically exhibit the appropriate mechanical, acoustical, and chemical properties suited for use with the invention.
  • well plates may be formed from polymeric material selected from the group consisting of polyethylenes, polypropylenes, polybutylenes, polystyrenes, cyclic olefins, combinations thereof, and copolymers of any of the foregoing.
  • Such polymers are generally inert to aqueous solutions and can be easily formed through casting, injection molding, extrusion, and other well-established processing techniques.
  • the invention also relates to reservoirs and well plates that exhibit a resistivity wherein the reservoir, the optional substrate, or both are comprised of a material that is at least partially polymeric and either has an electrical resistivity of no more than about 10 ohm-cm, has a surface electrical resistivity of no more than about 10 ohm/sq, or both.
  • polystyrene resin While most polymeric materials are insulators, conductive polymers are known in the art. For example, polythiophenes are a well-known class of conductive polymer and generally exhibit greater chemical stability than polyacetylene derivatives. Conductive polymer materials are extremely economical to produce and have been used commercially in the semiconductor field as containers for electrostatically sensitive materials. Relatively stable polythiophene derivatives include polyisothianapthene (PITN) and poly-3,4,ethylene dioxythiophene (PEDT), and a variety of related materials such as doped polypropylenes, are commercially available from RTP Company (Winona, Minnesota).
  • PITN polyisothianapthene
  • PEDT poly-3,4,ethylene dioxythiophene
  • an electrically conductive layer may be used to increase the conductivity of a reservoir.
  • a layer may be provided as a surface coating or incorporated within a reservoir to increase the reservoir's conductivity.
  • any part of an ordinary plastic well plate comprising an array of 96 substantially identical wells prone to accumulate uncontrolled electrostatic charge may be coated with a metallic coating.
  • metals such as aluminum, gold, silver, copper, platinum, palladium, or nickel may be selectively deposited on the upper, lower, interior, and/or exterior surface of an ordinary commercially available well plate.
  • plating technologies may be used to increase the thickness of the metallic coating.
  • nonmetallic coatings may be used as well.
  • known conductive ceramic coating materials include indium tin oxide and titanium nitride.
  • various forms of carbon e.g., carbon fibers, graphite, or acetylene black, may be applied as a surface coating on the reservoir.
  • a polymeric reservoir may contain an electrically conductive filler.
  • an electrically conductive filler Any of the materials suitable for forming the electrically conductive layer as discussed above may be used as a filler material.
  • carbon-filled plastics are well known in the art for electrostatic dissipation. Such carbon-filled plastics may be obtained from Minnesota Mining & Manufacturing Company Corporation (St. Paul, Minnesota) under the trademark Nelostat®. Such reservoirs may be formed using ordinary polymer processing techniques.
  • the acoustic radiation generator may have to be aligned with each reservoir during operation, discussed infra.
  • the center of each reservoir be located not more than about 1 centimeter, more preferably not more than about 1.5 millimeters, still more preferably not more than about 1 millimeter and optimally not more than about 0.5 millimeter, from a neighboring reservoir center. These dimensions tend to limit the size of the reservoirs to a maximum volume.
  • the reservoirs are constructed to contain typically no more than about 1 mL, preferably no more than about 100 ⁇ L, more preferably no more than about 10 ⁇ L, still more preferably no more than about 1 ⁇ L, and optimally no more than about 1 nL, of fluid.
  • the reservoirs may be either completely or partially filled with fluid.
  • fluid may occupy a volume of about 10 pL to about 100 nL.
  • each reservoir may be individually, efficiently, and systematically addressed.
  • arrays comprised of parallel rows of evenly spaced reservoirs are preferred.
  • each row contains the same number of reservoirs.
  • rectilinear arrays comprising X rows and Y columns of reservoirs are employed with the invention, wherein X and Y are each at least 2.
  • X may be greater than, equal to, or less than Y.
  • nonrectilinear arrays as well as other geometries may be employed. For example, hexagonal, spiral, or other types of arrays may be used.
  • the invention may be employed with irregular patterns of reservoirs, e.g., droplets randomly located on a flat substrate surface such as those associated with a CD-ROM format.
  • the invention may be used with reservoirs associated with microfluidic devices.
  • the invention may be used to dispense fluids of virtually any type and amount desired.
  • the fluid may be aqueous and/or nonaqueous.
  • fluids include, but are not limited to, aqueous fluids including waterier se and water-solvated ionic and non-ionic solutions; organic solvents; lipidic liquids; suspensions of immiscible fluids; and suspensions or slurries of solids in liquids.
  • fluids such as liquid metals, ceramic materials, and glasses may be used, as described in U.S. Patent Application Publication No. 20020140118.
  • the reservoir may contain a biomolecule, nucleotidic, peptidic, or otherwise.
  • the invention may be used in conjunction with dispensers for dispensing droplets of immiscible fluids, as described in U.S. Patent Application Publication Nos. 2002037375 and 20020155231, or to dispense droplets containing pharmaceutical agents, as discussed in U.S. Patent Application Publication No. 20020142049 and U.S. Patent Application Publication No. 20030012892 to et al.
  • any of a variety of focusing means may be employed to focus acoustic radiation so as to eject droplets from a reservoir.
  • one or more curved surfaces may be used to direct acoustic radiation to a focal point near a fluid surface.
  • Focusing means with a curved surface have been incorporated into the construction of commercially available acoustic transducers such as those manufactured by Panametrics Inc. (Waltham, Massachusetts).
  • Fresnel lenses are known in the art for directing acoustic energy at a predetermined focal distance from an object plane. See, e.g., U.S. Patent No.
  • Fresnel lenses may have a radial phase profile that diffracts a substantial portion of acoustic energy into a predetermined diffraction order at diffraction angles that vary radially with respect to the lens.
  • the diffraction angles should be selected to focus the acoustic energy within the diffraction order on a desired object plane.
  • acoustic focusing means exhibiting a variety of F-numbers may be employed with the invention. As discussed in U.S. Patent No.
  • the focusing means suitable for use with the invention typically exhibits an F-number of at least about 1.
  • the focusing means exhibits an F-number of at least about 2.
  • a preferred approach is to acoustically couple the ejector to the reservoir without contacting any portion of the ejector, e.g., the focusing means, with the fluids to be . ejected.
  • a positioning means is provided for positioning the ejector in controlled and repeatable acoustic coupling with each of the fluids in the reservoirs to eject droplets therefrom without submerging the ejector therein. This typically involves direct or indirect contact between the ejector and the external surface of each reservoir.
  • the direct contact be wholly conformal to ensure efficient acoustic energy transfer. That is, the ejector and the reservoir should have corresponding surfaces adapted for mating contact. Thus, if acoustic coupling is achieved between the ejector and reservoir through the focusing means, it is desirable for the reservoir to have an outside surface that corresponds to the surface profile of the focusing means. Without conformal contact, efficiency and accuracy of acoustic energy transfer may be compromised. In addition, since many focusing means have a curved surface, the direct contact approach may necessitate the use of reservoirs having a specially formed inverse surface.
  • an acoustic coupling medium may be interposed between the reservoir and ejector.
  • the acoustic coupling medium is a fluid.
  • the acoustic coupling medium is preferably an acoustically homogeneous material that is substantially free of material having different acoustic properties than the fluid medium itself.
  • the acoustic coupling medium be comprised of a material having acoustic properties that facilitate the transmission of acoustic radiation without significant attenuation in acoustic pressure and intensity.
  • the acoustic impedance of the coupling medium should facilitate the transfer of energy from the coupling medium into the reservoir.
  • An aqueous fluid such as waterier se, may be employed as an acoustic coupling medium.
  • Ionic additives e.g., salts, may sometimes be added to the coupling medium to increase the conductivity of the coupling medium, thereby facilitating discharge of any charge accumulated by the reservoir.
  • a single ejector is preferred, although the inventive device may include a plurality of ejectors.
  • the means for positioning the ejector may be adapted to provide relative motion between the ejector and reservoirs.
  • the positioning means should allow for the ejector to move from one reservoir to another quickly and in a controlled manner, thereby allowing fast and controlled scanning of the reservoirs to effect droplet ejection therefrom.
  • various means for positioning the ejector in acoustic coupling relationship to the reservoir are generally known in the art and may involve, e.g., devices that provide movement having one, two, three, four, five, six, or more degrees of freedom.
  • the ejector may be movable in a row- wise direction and/or in a direction perpendicular to both the rows and columns.
  • the rate at which fluid droplets can be delivered is related to the efficiency of fluid delivery.
  • Current positioning technology allows for the ejector positioning means to move from one reservoir to another quickly and in a controlled manner, thereby allowing fast and controlled ejection of different fluid samples. That is, current commercially available technology allows the ejector to be moved from one reservoir to another, with repeatable and controlled acoustic coupling at each reservoir, in less than about 0.1 second for high performance positioning means and in less than about 1 second for ordinary positioning means.
  • a custom designed system will allow the ejector to be moved from one reservoir to another with repeatable and controlled acoustic coupling in less than about 0.001 second.
  • the invention also enables rapid ejection of droplets from one or more reservoirs, e.g., at a rate of at least about 1,000,000 droplets per minute from the same reservoir, and at a rate of at least about 100,000 drops per minute from different reservoirs, assuming that the droplet size does not exceed about 10 ⁇ m in diameter.
  • the droplet generation rate is a function of drop size, viscosity, surface tension, and other fluid properties. In general, the droplet generation rate increases with decreasing droplet diameter, and 1,000,000 droplets per minute is achievable for most aqueous fluid drops under about 10 ⁇ m in diameter.
  • the invention may be used in any context where precise placement of a fluid droplet is desirable or necessary.
  • the invention may be employed to improve accuracy and precision associated with nozzleless acoustic ejection.
  • acoustic ejection technology may be used to form biomolecular arrays.
  • acoustic ejection technology may be employed to format a plurality of fluids, e.g., to transfer fluids from odd-sized bulk containers to wells of a standardized well plate or to transfer fluids from one well plate to another.
  • focused acoustic radiation may serve to eject a droplet of fluid from a reservoir into any sample vessel for processing and/or analyzing a sample molecule, e.g., into a sample introduction interface of a mass spectrometer, an inlet opening that provides access to the interior region of a capillary, or an inlet port of a microfluidic device.
  • the invention may be used to eject droplets of analysis-enhancing fluid on a sample surface in order to prepare the sample for analysis, e.g., for MALDI or SELDI-type analysis.
  • uncontrolled electrostatic charge may be accumulated by a substrate onto which droplets are dispensed. Such charge may also have a detrimental influence on the trajectory and/or volume of the dispensed droplets.
  • construction considerations for such substrates are similar to those associated with reservoirs, as discussed above.
  • the substrate may exhibit a relatively high electrical conductivity for ease in grounding.
  • the materials and techniques suitable for use in forming the reservoir may also be used with the substrate. In some instances, a single means for reducing uncontrolled charge may be used for both the reservoir and substrate.
  • the invention may also employ a positioning means for positioning the substrate.
  • a positioning means for positioning the substrate.
  • pulse motion involves the discrete steps of moving an ejector into position, emitting acoustic energy, and moving the ejector to the next position; again, using a high performance positioning means with such a method allows repeatable and controlled acoustic coupling at each reservoir in less than 0.1 second.
  • a continuous motion design moves the ejector and the reservoirs continuously, although not necessarily at the same speed, and provides for ejection during movement. Since the pulse width is very short, this type of process enables over 10 Hz reservoir transitions, and even over 1000 Hz reservoir transitions.
  • relative motion between the reservoirs and the acoustic generator can be achieved by moving the reservoirs while holding the generator still, by moving the reservoirs while holding the generator still, or by moving the generator and the reservoirs at different velocities. Similar engineering considerations are applicable to the substrate positioning means.
  • the various components of the device may require individual control or synchronization to direct droplets onto designated sites on a substrate surface.
  • the ejector positioning means may be adapted to eject droplets from each reservoir in a predetermined sequence associated with an array of designated sites on the substrate surface.
  • Any positioning means of the present invention may be constructed from, e.g., levers, pulleys, gears, a combination thereof, or other mechanical means known to one of ordinary skill in the art.
  • acoustic energy may be used to assess the contents of a plurality of reservoirs.
  • acoustic assessment technologies are described U.S. Patent Application Publication Nos. 20030101819 and 20030150257, each to Mutz et al.
  • the acoustic radiation generator may be used in combination with an analyzer for analyzing a characteristic of acoustic radiation generated by the generator and transmitted through the reservoir. By placing the analyzer in radiation receiving relationship to the acoustic radiation generator, the acoustic radiation having interacted with the contents of the reservoir may be analyzed.
  • Such acoustic assessment of the contents of one or more reservoirs may enhance the accuracy and precision in dispensing fluids therefrom.
  • a means for reducing uncontrolled electrostatic charge is employed so that any dispensed droplet exhibits a volume and/or trajectory that conform to a predetermined volume and/or trajectory.
  • the means for reducing uncontrolled electrostatic charge is selected according to the location, amount, and type of static electricity to be eliminated.
  • the means for reducing uncontrolled electrostatic charge must be constructed according to the construction of the reservoir.
  • the means for reducing electrostatic charge may be constructed accordingly.
  • any effort to eliminate uncontrolled electrostatic charge may ensure that a droplet dispensed from the reservoir has a volume that does not deviate from the predetermined volume by more than about 10 %.
  • the droplet volume does not deviate from the predetermined volume by more than about 5 %.
  • the volume does not deviate from the predetermined volume by more than about 2 %.
  • the trajectory of the droplet dispensed from the reservoir will typically not deviate from the predetermined trajectory by more than about 5°.
  • the trajectory does not deviate from the predetermined trajectory by more than about 1°.
  • the trajectory does not deviate from the predetermined trajectory by more than about 0.5°.
  • electrostatic control techniques are known in the art and are suited for use with the present invention. Such techniques typically involve either addition or removal of electrons from the item that has accumulated uncontrolled electrostatic charge. On occasion, though, positive ions may be added or removed from the item. In general, electrostatic charge can be removed through grounding, induction, ionization, or a combination thereof. Such electrostatic charge neutralization may be effected immediately before or during the dispensation of a droplet.
  • uncontrolled electrostatic charge may be eliminated from an item through grounding, i.e., connecting the item via a conductor to an effectively infinite source of charge.
  • Grounding is particularly suited for instances in which electrostatic charge is located in an ungrounded but highly conductive item. In such a case, the entire item may be neutralized when it is connected to ground at a single point.
  • items constructed from a material having a volume electrical resistivity of no more than about 10 4 ohm-cm and/or a surface electrical resistivity of no more than about 10 5 ohm/sq may be used.
  • the electrical resistivity is no more than about 10 3 ohm-cm and/or the surface electrical resistivity is no more than about 10 ohm/sq.
  • neutralization of the entire item may require the establishment of more than a single-point contact. In some instances, neutralization of an item may be achieved by providing the item with intermittent or sustained contact with an electrically conductive solid material.
  • Removing or neutralizing electrostatic charge by induction is a time-tested method suitable for use with any nonconductive material, insulated material, or ungrounded conductive material.
  • Induction requires the use of an electrically conductive induction member that operates in a manner similar to the operation of a lightning rod.
  • a grounded induction member such as tinsel or a brush, is placed in close proximity, e.g., about 0.5 cm to about 1.0 cm, to the surface of the material to be neutralized. If the electrostatic charge on the material reaches or exceeds a threshold level, e.g., at least several thousand volts, the energy concentrated on the ends of the induction member will induce ionization.
  • a threshold level e.g., at least several thousand volts
  • induction may not reduce or neutralize static electricity to the ground potential level.
  • an ungrounded induction member will remove charge for a short period of time only. Eventually the induction member will self charge and stop working when the electric field between the ends and the charged surface is reduced to a level that cannot support ionization.
  • passive static control devices relying solely on induction tend to leave a residual charge.
  • Ionization techniques typically involve the production of both positive and negative ions to be attracted by the material to be neutralized. This may be achieved by generating an alternating electric field between a sharp point in close proximity to a grounded shield or casing. As the extremes of potential difference are reached, the air between the sharp point and the grounded casing is broken down. As a result, positive and negative ions are generated. In other words, half of the cycle is utilized to generate negative ions and the other half is utilized to generate positive ions. When a 60 Hz unit is employed, the polarity of ionization is changed every 1/120 of a second. If the material to be neutralized is positively charged, it will immediately absorb negative ions and repel the positive ions into space. Conversely, if the material to be neutralized is negatively charged, it will absorb the positive ions and repel the negative ions. When the material becomes neutralized, there is no longer electrostatic attraction and the material will cease to absorb ions.
  • Other equipment may also be used to generate ionized air for electrostatic neutralization.
  • Nuclear-powered ionizers are known in the art.
  • Polonium 210 isotopes may be used to generate ions. Since Polonium has a half-life of only 138 days, such ionizers continually lose their strength and must be replaced annually.
  • electromagnetic radiation sources may be used to eliminate electrostatic charge. In some instances, such electromagnetic sources employ an ultraviolet radiation generator.
  • surface conductivity of an item may be increased through the use of additives such anti-static sprays.
  • An ordinary anti-static spray is comprised of a surfactant diluted in a solvent.
  • a fire retardant may be added to counter the flammability of the solvent.
  • the fire retardant and solvents evaporate, leaving a conductive coating on the surface of the material.
  • the plastic has now become conductive and as long as this coating is not disturbed, it will be difficult to generate static electricity in this material.
  • neutralization of an item may involve establishing intermittent or prolonged contacting of the item with a liquid and/or electrostatic-charge-reducing fluid.
  • the acoustic coupling medium may be comprised of an electrostatic-charge-reducing fluid.
  • the invention preferably involves dispensing one or more droplets in the absence of any electrostatic charge or electric field that alters the trajectory and/or size of dispensed droplets.
  • production of a droplet of appropriate direction, volume, and velocity is accompanied by the production of a secondary or satellite droplet that should not be deposited onto the droplet-receiving surface.
  • Using an electric field may accelerate both drops onto a receiving surface.
  • electric fields may adversely interfere with droplet formation so as to result in difficulty in controlling droplet size.
  • FIG. 1 illustrates an exemplary focused acoustic ejection device suitable for use with the invention, in simplified cross-sectional view.
  • the device 11 includes a plurality of reservoirs, i.e., at least two reservoirs— a first reservoir indicated at 13 and a second reservoir indicated at 15.
  • Each reservoir may contain a combination of two or more immiscible fluids, and the individual fluids, and the fluid combinations in the different reservoirs may be the same or different.
  • reservoir 13 contains fluid 14
  • reservoir 15 contains fluid 16.
  • Fluids 14 and 16 have fluid surfaces respectively indicated at 14S and 16S.
  • the reservoirs are of substantially identical construction so as to be substantially acoustically indistinguishable, but identical construction is not a requirement.
  • the reservoirs are shown as separate removable components but may, if desired, be fixed within a plate or other substrate.
  • Each of the reservoirs 13 and 15 is axially symmetric as shown, having vertical walls 13W and 15W extending upward from circular reservoir bases 13B and 15B and terminating at openings 13O and 15O, respectively, although other reservoir shapes may be used.
  • the material and thickness of each reservoir base should be such that acoustic radiation may be transmitted therethrough and into the fluid contained within the reservoirs.
  • the device also includes an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation, and a focusing means 37 for focusing the acoustic radiation at a focal point near the fluid surface from which a droplet is to be ejected, wherein the focal point is selected so as to result in droplet ejection.
  • the focal point may be in any layer.
  • the focal point may be in the upper fluid layer or in the lower fluid layer, e.g., just below the interface therebetween. As shown in FIG.
  • the focusing means 37 may comprise a single solid piece having a concave surface 39 for focusing acoustic radiation, but the focusing means may be constructed in other ways as discussed below.
  • the acoustic ejector 33 is thus adapted to generate and focus acoustic radiation so as to eject a droplet of fluid from each of the fluid surfaces 14S and 16S when acoustically coupled to reservoirs 13 and 15, respectively.
  • the acoustic radiation generator 35 and the focusing means 37 may function as a single unit controlled by a single controller, or they may be independently controlled, depending on the desired performance of the device.
  • single ejector designs are preferred over multiple ejector designs, because accuracy of droplet placement, as well as consistency in droplet size and velocity, are more easily achieved with a single ejector.
  • acoustic coupling is achieved between the ejector and each of the reservoirs through indirect contact.
  • an acoustic coupling medium 25 is placed between the ejector 33 and the base 13B of reservoir 13, with the ejector and reservoir located at a predetermined distance from each other.
  • the acoustic coupling medium 25 is introduced from a coupling medium source 27 via dispenser 29.
  • an optional collector 47 is employed to collect coupling medium that may drip from the lower surface of either reservoir. As the collector 47 is depicted as containing the coupling medium source 27, it is evident that the coupling medium may be reused.
  • the coupling medium source 27 and dispenser 29 serve as a means for reducing uncontrolled electrostatic charge from the reservoirs.
  • each reservoir 13 and 15 of the device is filled with different fluids, as explained above.
  • the acoustic ejector 33 is positionable by means of ejector positioning means 61, shown below reservoir 13, in order to achieve acoustic coupling between the ejector and the reservoir through acoustic coupling medium 25.
  • a substrate 53 may be positioned above and in proximity to the first reservoir 13 such that one surface of the substrate, shown in FIG. 1 as underside surface 53S, faces the reservoir and is substantially parallel to the surface 14S of the fluid 14 therein.
  • the substrate 53 is held by substrate positioning means 65, which, as shown, is grounded.
  • the substrate 53 is comprised of a conductive material
  • the substrate 53 is grounded as well.
  • the acoustic radiation generator 35 is activated to produce acoustic radiation that is directed by the focusing means 37 to a focal point 14P near the fluid surface 14S of the first reservoir.
  • droplet 14D is ejected from the fluid surface 14S, optionally onto a particular site (typically although not necessarily, a pre-selected, or "predetermined" site) on the underside surface 53S of the substrate.
  • the ejected droplet may be retained on the substrate surface by solidifying thereon after contact; in such an embodiment, it is necessary to maintain the substrate surface at a low temperature, i.e., at a temperature that results in droplet solidification after contact.
  • a molecular moiety within the droplet attaches to the substrate surface after contact, through adsorption, physical immobilization, or covalent binding.
  • a substrate positioning means 65 may be used to reposition the substrate 53 (if used) over reservoir 15 in order to receive a droplet therefrom at a second site.
  • FIG. IB also shows that the ejector 33 has been repositioned by the ejector positioning means 61 below reservoir 15 and in acoustically coupled relationship thereto by virtue of acoustic coupling medium 25.
  • the acoustic radiation generator 35 of ejector 33 is activated to produce acoustic radiation that is then directed by focusing means 37 to a focal point 16P within the reservoir fluids in reservoir 15, thereby ejecting droplet 15D, optionally onto the substrate.
  • the inventive device may be used to eject a plurality of droplets from reservoirs in order to form a pattern, e.g., an array, on the substrate surface 53S. It should be similarly evident that the device may be adapted to eject a plurality of droplets from one or more reservoirs onto the same site of the substrate surface. Furthermore, the ejection of a plurality of droplets may involve one or more ejectors. In some instances, the droplets are ejected successively from one or more reservoirs. In other instances, droplets are ejected simultaneously from different reservoirs.
  • the invention may be used with a single reservoir as well to improve the accuracy of droplet dispensation therefrom into an inlet opening of a sample vessel.
  • Axially symmetric and grounded capillary 53 having an inlet opening 53O disposed on a terminus 53S thereof is provided as a sample vessel. Due to the axial symmetry of the capillary 53, the inlet opening 53O has a circular cross section. As such, the opening has a limiting dimension equal to its diameter.
  • a hemispherical volume of fluid 14 on a substantially flat surface 13S of a substrate 13 serves a reservoir. As shown, the substrate 13 is grounded so that it does not have any uncontrolled electrostatic charge.
  • the shape of fluid 14 is a function of the sample wetting properties with respect to the substrate surface 13S. Thus, the shape can be modified with any of a number of surface modification techniques.
  • an ejector 33 is provided comprising an acoustic radiation generator 35 for generating radiation, and a focusing means 37 for directing the radiation at a focal point 14P near the surface 14S of the fluid 14. The ejector 33 is shown in acoustic coupling relationship to the substrate 13 through coupling fluid 25.
  • the invention may be suitable for use with any of the performance enhancing features associated with acoustic technologies such those described in U.S. Patent Application Publication No. 20030230344 to Ellson et al. relating to acoustic control of the composition and/or volume of fluid in a reservoir.
  • the invention may be used in a number of contexts such as handling pathogenic fluids and manipulating cells and particles (see U.S. Patent Application Publication Nos. 20020090720 and 20020094582).
  • a single acoustic radiation generator when used for ejecting and optionally assessing the contents of a plurality of reservoirs, the generator may be placed in acoustic coupling relationship in rapid succession to each of the reservoirs via the acoustic coupling fluid. Accordingly, the generator, the reservoirs, or both must be rapidly displaced with respect to each other for high-throughput techniques. Such rapid movement may cause uncontrolled flow of the acoustic coupling fluid. As a result, conformal contact between the acoustic generator and the reservoirs may not be achieved, thereby compromising the performance of the device. In some instances, uncontrolled acoustic fluid flow may result in the contamination of the reservoir contents, presence of sound-reflecting bubbles in the acoustic path, and/or degradation of device components.
  • an acoustic coupling medium is comprised of an electrostatic-charge-reducing fluid
  • nonconforrhal contact of the coupling medium to the reservoir may interfere with the dissipation of uncontrolled electrostatic charge on the reservoir.
  • a means may be provided for eliminating uncontrolled flow of the acoustic coupling fluid at the exterior surface as a result of movement of the acoustic radiation generator. Such means may serve to facilitate and improve control over the elimination of electrostatic charge.
  • FIG. 3 depicts an acoustic device prone to such problems in simplified cross- sectional view.
  • the device is shown in operation to form a biomolecular array bound to a substrate.
  • the device 11 is generally similar to that depicted in FIG. 1 and includes two reservoirs, with a first reservoir indicated at 13 and a second reservoir indicated at 15. As shown, the first reservoir 13 contains a first fluid 14 and the second reservoir 15 contains a second fluid 16.
  • Fluids 14 and 16 each have a fluid surface respectively indicated at 14S and 16S. As depicted, fluids 14 and 16 are of differing volumes and heights. That is, the distance between surface 14S and base 13B is greater than the distance between surface 16S and base 15B.
  • the device also includes an acoustic radiation generator 35 that contains a transducer 36, e.g., a piezoelectric element, commonly shared by an analyzer.
  • a combination unit 38 is provided that both serves as a controller and a component of an analyzer. Operating as a controller, the combination unit 38 provides the piezoelectric element 36 with electrical energy that is converted into mechanical and acoustic energy. Operating as a component of an analyzer, the combination unit receives and analyzes electrical signals from the transducer. The electrical signals are produced as a result of the absorption and conversion of mechanical and acoustic energy by the transducer.
  • the focusing means 37 is comprised of a single solid piece having a concave surface 39 for focusing acoustic radiation.
  • the acoustic ejector 33 is thus adapted to generate and focus acoustic radiation so as to eject a droplet of fluid from each of the fluid surfaces 17 and 19 when acoustically coupled to reservoirs 13 and 15, and thus to fluids 14 and 16, respectively.
  • the acoustic radiation generator 35 and the focusing means 37 function as a single unit controlled by a single controller.
  • a dispenser 29 places an acoustic coupling fluid 25 between the ejector 33 and the base 13B of reservoir 13, with the ejector placed at a predetermined distance from each the reservoir by positioning means 61.
  • the dispenser 29 dispenses sufficient coupling fluid 25 so that the fluid established conformal contact between the concave surface 39 and base 13B.
  • the acoustic radiation generator 35 is activated to produce acoustic radiation that is directed toward a free fluid surface 14S of the first reservoir.
  • the acoustic radiation will then travel in a generally upward direction toward the free fluid surface 14S.
  • the acoustic radiation will be reflected.
  • substrate By determining the time it takes for the acoustic radiation to be reflected by the fluid surface back to the acoustic radiation generator, and then correlating that time with the speed of sound in the fluid, the distance-and thus the fluid height-may be calculated. [0103] In order to form a biomolecular array on a substrate using the device, substrate
  • the ejector 33 is moved toward to the reservoir 13 to ensure that the focal point of the ejection acoustic wave is near the fluid surface 14S, where desired. That is, the ejector 33 is moved positively along axis Z. As a result, acoustic coupling fluid 25 is displaced through uncontrollable flow. When movement of the ejector is at a high velocity, the acoustic coupling fluid may be squirted or sprayed in a direction perpendicular to axis Z.
  • the acoustic radiation generator 35 is activated to produce acoustic radiation that is directed by the focusing means 37 to a focal point 14P near the fluid surface 14S of the first reservoir. That is, an ejection acoustic wave having a focal point near the fluid surface is generated in order to eject at least one droplet of the fluid. As a result, droplet 14D is ejected from the fluid surface 14S onto a designated site on the underside surface 51 of the substrate.
  • FIG. 3C a substrate positioning means 65 repositions the substrate 53 over reservoir 15 in order to receive a droplet therefrom at a second designated site.
  • FIG. IC also shows that the ejector 33 has been repositioned by the ejector positioning means 61 below reservoir 15 and in acoustically coupled relationship thereto by virtue of acoustic coupling fluid 25.
  • the dispenser 29 dispenses sufficient coupling fluid 25 so that the fluid establishes conformal contact between the concave surface 39 and base 15B.
  • the acoustic radiation generator 35 of ejector 33 is activated to produce low energy acoustic radiation to assess the height of fluid 16 in reservoir 15 and to determine whether and/or how to eject fluid from the reservoir.
  • acoustic coupling fluid 25 flows uncontrollably so that it no longer conforms to surface 39 and base 15B.
  • bubbles will form within the acoustic coupling fluid. For example, air bubbles may be sucked into fluid. Under extreme circumstances, bubbles may be formed as a result of cavitation.
  • any droplet 16D ejected from reservoir 15 toward substrate 53 may be misdirected due to the lack of conformal contact.
  • another embodiment of the invention provides an acoustic device as described above that also includes a means for delivering an acoustic coupling fluid to an exterior surface of the reservoir and a means for eliminating uncontrolled flow of the acoustic coupling fluid at the exterior surface as a result of movement of the acoustic radiation generator.
  • any of a number of different means may be used to deliver the acoustic coupling fluid to the exterior surface of the reservoir, such means typically includes a source of the acoustic coupling fluid in fluid communication with a nozzle having an outlet that opens toward the exterior surface of the reservoir.
  • the acoustic coupling fluid is comprised of water.
  • fluids similar to water may be used as well.
  • the acoustic coupling medium may be comprised of a fluid that exhibits an attenuation coefficient for acoustic radiation of a selected frequency similar to that of water. The selected frequency is typically the operating frequency of the device.
  • the coupling fluid exhibits an attenuation coefficient for acoustic radiation of a selected frequency that differs from the attenuation coefficient of water at the same frequency by no more than about 10%.
  • the difference in attenuation coefficient is no more than about 5%.
  • the difference in attenuation coefficient is no more than about 1%.
  • the acoustic coupling fluid is typically directed to flow from the source to the outlet at a rate sufficient for the acoustic coupling fluid to establish conformal contact with the exterior surface of the reservoir.
  • the device may include a collector as well as a means for positioning the nozzle.
  • the collector is placed in fluid-receiving relationship to the exterior surface of the reservoir so as to collect excess acoustic coupling fluid flowing therefrom.
  • the nozzle may be placed directly below the exterior surface of the reservoir such that acoustic coupling fluid emerging from the nozzle is directed upward for conformal contact with the exterior surface of the reservoir.
  • the nozzle may be located within the collector.
  • the nozzle is typically placed no closer than a predetermined distance from the exterior surface of the reservoir so as to avoid contact between the nozzle and the surface.
  • some embodiments allow acoustic radiation is propagated through the acoustic coupling fluid in the nozzle and the exterior surface into the reservoir.
  • a particularly useful design allows the nozzle and the acoustic radiation generator to move along the same axis extending from the exterior surface of the reservoir. Typically, the axis is perpendicular to the exterior surface.
  • FIG. 4 depicts an exemplary acoustic device having a nozzle and collector as described above.
  • a single reservoir 13 containing a fluid 14 having a fluid surface indicated at 14S.
  • Reservoir 13 has a base indicated at 13B and an opening indicated at 13O.
  • Dispenser 29 provided is comprised of a nozzle 30 that terminates upwardly at an outlet 32 directed toward the reservoir base 13B and downwardly at a pump 34 for pumping acoustic coupling fluid 25 upwardly through the nozzle 30.
  • an acoustic ejector 33 Located within the nozzle 30 is an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation and a focusing means 37 for focusing the acoustic radiation at a focal point within the fluid 14P from which a droplet is to be ejected, near the fluid surface 14S.
  • Positioning means 61 serves to controllably move ejector 33 within nozzle 30 along axis Z.
  • the device also includes a collector 31 for collecting coupling fluid that flows from base 13B. As shown, the nozzle 30 is located within the collector 31. Located at the bottom of the collector 31 and in fluid communication with the pump 34 is a source 27 of acoustic coupling fluid.
  • positioning means 70 positions the dispenser 29 at predetermined distance to the reservoir base 13B.
  • the pump 34 draws acoustic coupling fluid from the source 27 and forces the acoustic coupling fluid upward through the nozzle 30.
  • the flow of acoustic coupling emerging from outlet 32 is typically maintained at constant rate and sufficiently high to allow the coupling fluid to establish and maintain conformal contact with reservoir base 13B. After contact with reservoir base 13B, acoustic coupling fluid falls back down into collector 31, where the coupling fluid redirected toward source 27 and pump 34 for reuse.
  • the acoustic coupling fluid 25 between the ejector 33 and the base 13B allows for acoustic radiation generated by the generator 35 to be transmitted therethrough.
  • acoustic radiation will then travel in a generally upward direction, through base 13B and fluid 14 toward the free fluid surface 14S.
  • the acoustic radiation reflected by free surface 14S may then be analyzed. If needed to ensure that the acoustic radiation is focused near the fluid surface 14S to eject a droplet therefrom, positioning means in the form of telescoping rod 61 may be employed to move ejector 33 to an appropriate location within nozzle 30.
  • the rod 61 may be adapted to elongate in a telescoping manner within the nozzle to move ejector 33 toward the outlet 32. Similarly, the ejector 33 is moved toward pump 34 when rod 61 is retracted. In any case, the ejector 33 may be maintained at a fixed distance from the fluid surface 14S so as to ensure that the acoustic radiation remains focused near the fluid surface 14S as the fluid level in the reservoir 13 is lowered due to the ejection of droplets therefrom.
  • a number of different designs and mechanisms may be used as a means for eliminating uncontrolled flow of the acoustic coupling fluid.
  • uncontrolled fluid flow may be avoided simply by immobilizing the relative positions of the reservoir 13 and the nozzle 30 and maintaining fluid flow from outlet 32 at a constant rate.
  • any movement of ejector 33 within nozzle 30, particularly rapid movement may disturb the rate of fluid flow from outlet 32, particularly when the pump 34 moves acoustic fluid at a constant rate.
  • the rate of fluid flow emerging from outlet 32 will tend to increase temporarily as rod 61 displaces as coupling fluid within the nozzle 30.
  • movement of ejector 33 downward toward pump 34 will cause the rate of fluid flow emerging from outlet 32 to decrease.
  • means for eliminating uncontrolled coupling fluid flow from outlet 32 may serve to maintain the fluid pressure at outlet 32 at a constant level.
  • the means for positioning the nozzle and the means for positioning the generator may be synchronized to maintain flow of acoustic coupling fluid from the nozzle at a constant rate, thereby serving as the means for eliminating uncontrolled flow.
  • a displacement member that maintains the acoustic coupling fluid at a constant volume within the nozzle may be used in response any movement of the acoustic radiation generator within the nozzle.
  • Such displacement members may be selected from pistons, diaphragm, combinations thereof, and other mechanisms.
  • the displacement member may be at least partially located within the nozzle.
  • the displacement member may be at least partially located external to the nozzle in a chamber that fluidly communicates with the nozzle.
  • a flow rate regulator may be advantageously used to adjust the flow rate of the acoustic coupling fluid from the source to the outlet according to movement of the acoustic radiation generator within the nozzle.
  • an adjustable valve may be provided downstream from the source and upstream from the outlet to adjust the flow rate of the acoustic coupling fluid.
  • Flow rate regulator technology is well known in the art and one of ordinary skill should be able to adapt the device to incorporate such regulators.
  • FIG. 5 depicts an exemplary acoustic device that employs a stationary opposing piston design to maintain coupling fluid flow at a constant rate from a nozzle outlet. The opposing piston design operates by maintaining the acoustic coupling fluid at a constant volume within the nozzle.
  • Dispenser 29 provided is comprised of a nozzle 30 that terminates upwardly at an outlet 32 directed upwardly for delivering acoustic coupling fluid 25 to the exterior surface of a reservoir (not shown).
  • a nozzle 30 Located within the nozzle 30 is an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation and a focusing means 37 for focusing the acoustic radiation.
  • Positioning means in the form of a platform 61 serves to move ejector 33 within nozzle 30 along axis Z in a controlled manner.
  • a stationary piston 72 that extends through the nozzle 30 and into a corresponding opening 74 in platform 61. As depicted, the volume of acoustic coupling fluid 25 within the nozzle remains constant as ejector 33 is moved along axis Z as long as piston 72 extends through opening 74.
  • a means other than a nozzle may be used to deliver acoustic coupling fluid to the exterior surface of a reservoir.
  • a container may be sealed against the reservoir and filled with the acoustic coupling fluid such that the acoustic coupling fluid is in conformal contact with the exterior surface of the reservoir.
  • the acoustic radiation generator may be movable within the container.
  • FIG. 6 depicts an acoustic device similar to that depicted in FIG. 3 with some notable differences relating to the means for eliminating uncontrolled coupling fluid flow.
  • the device 11 includes two attached reservoirs provided in the form of wells 13 and 15 of a well plate 12. The wells 13 and 15 share a common underside surface 12B that is substantially planar.
  • the device of FIG. 6 also includes an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation and a focusing means 37 for focusing the acoustic radiation at a focal point within the fluid from which a droplet is to be ejected, near the fluid surface.
  • a positioning means 61 serves to couple the ejector 33 successively to each of the wells.
  • the dispenser is replaced with a container
  • acoustic ejector 33 and positioning means 61 are both sealed within the container 29 and submerged in coupling fluid 25. Because the volume of the coupling fluid remains 25 unaltered irrespective of the movement and or positioning of the ejector 33 within the container 29, uncontrolled flow of the acoustic coupling fluid at the exterior surface as a result of movement of the acoustic radiation generator is eliminated.
  • any of the above-described means for eliminating uncontrolled coupling fluid flow associated with the nozzle may be used with the container as well.
  • a method for transmitting acoustic radiation into a reservoir involves simultaneously delivering an acoustic coupling fluid to an exterior surface of a reservoir adapted to contain a fluid and positioning an acoustic radiation generator for generating acoustic radiation in acoustic coupling relationship via the acoustic coupling fluid to the reservoir. This is carried out in a manner that avoids uncontrolled flow of the acoustic coupling fluid at the exterior surface. Once the acoustic radiation generator is in position, it is activated so as to generate and transmit acoustic radiation through the exterior surface and into any fluid contained in the reservoir.
  • positioning means any of a number of positioning means known in the art may be used with the invention.
  • Such positioning means may be constructed from, e.g., levers, pulleys, gears, a combination thereof, or other mechanical means known to one of ordinary skill in the art.
  • positioning means may be used to move items such as the reservoir, the acoustic generator, the coupling fluid delivering means, or a combination thereof, to provide relative motion therebetween.
  • relative motion may be provided by holding any one or a combination of the items in a fixed position while the allowing the positioning means to move the remaining items.
  • acoustic radiation generators such as transducer assemblies may be used as well. That is, linear acoustic arrays, curvilinear acoustic arrays, annular acoustic arrays, phased acoustic arrays, and other transducer assemblies may be used in conjunction with the invention as well.
  • acoustic detectors like acoustic generators, may be used in conjunction with acoustic coupling fluids, those of ordinary skill in the art will be able to substitute acoustic detectors in place of acoustic generators in certain applications.
  • a solution containing 70% by volume dimethylsulfoxide and 30% by volume water was placed within each well of a polystyrene well plate containing 384 substantially identical wells.
  • An acoustic ejector having an F2 lens that served to focus acoustic radiation was placed in acoustic coupling relationship successively with each reservoir in substantially the same manner. Without removing uncontrolled electrostatic charge from the well plate, acoustic radiation having a frequency of 10 MHz was directed by the F2 lens into each reservoir so as to eject at least one droplet from each well.
  • secondary or satellite droplets were produced in addition to the primary droplets. The primary droplets exhibited a volume variation of over 25% as well as variations in trajectory.
  • Each well of the same polystyrene well plate described in Example 1 was again filled with a solution containing 70% by volume dimethylsulfoxide and 30% by volume water.
  • uncontrolled electrostatic charge was removed from the well plate using an ionizer before the acoustic ejector was placed in acoustic coupling relationship successively with each reservoir.
  • Acoustic radiation of having a frequency of 10 MHz was again directed by the F2 lens into each reservoir so as to eject at least one droplet from each well.
  • No secondary or satellite droplets were produced.
  • the primary droplets exhibited a volume variation of less than about 2%. No variations in the trajectory of the droplets were observed.

Abstract

Devices and methods are provided for reducing uncontrolled electrostatic charge that can alter the volume and/or trajectory of a droplet, which may be ejected through the application of focused acoustic radiation. Also provided are reservoirs and substrates, e.g., well plates formed from a material that is at least partially nonmetallic or polymeric and either has an electrical resistivity of no more than about 1011 ohm-cm, has a surface electrical resistivity of no more than about 1012 ohm/sq, or both.

Description

DRQPLET DISPENSATION FROM A RESERVOIR WITH REDUCTION IN UNCONTROLLED ELECTROSTATIC CHARGE
TECHNICAL FIELD
[0001] This invention relates generally to devices and methods for accurately dispensing a droplet from a reservoir, optionally toward a substrate, wherein the volume and/or trajectory of the droplet do not substantially deviate from a predetermined volume and/or trajectory. More particularly, the invention relates to devices and methods for reducing the uncontrolled electrostatic charges that can alter the volume and/or trajectory of a droplet, which is typically ejected through the application of focused acoustic radiation.
BACKGROUND ART
[0002] There exists a need in pharmaceutical, biotechnological, medical, and other industries to be able to quickly screen, identify, analyze, and/or process large numbers or varieties of fluids. As a result, much attention has been focused on developing efficient, precise, and accurate fluid handling methods. High-speed combinatorial methods often involve the use of array technologies that require accurate dispensing of fluids. For example, automated robotic systems have been used in combination with precise registration technologies to dispense reagents through automated pick-and-place ("suck-and-spit") fluid handling systems. Similarly, some efforts have been directed to adapting printing technologies, particularly inkjet printing technologies, to form biomolecular arrays. For example, U.S. Patent No. 6,015,880 to Baldeschwieler et al. is directed to array preparation using multistep in situ synthesis. Such synthesis may involve using inkjet technology to dispense reagent-containing droplets to a locus on a surface chemically prepared to permit covalent attachment of the reagent.
[0003] Such conventional fluid handling systems, however, exhibit certain inherent disadvantages. For example, most fluid handling systems presently in use require that contact be established between the fluid to be transferred and an associated solid surface on the transferring device. Such contact typically results in surface wetting that causes unavoidable fluid waste, a notable drawback when the fluid to be transferred is rare and/or expensive. When fluid dispensing systems are constructed using networks of tubing or other fluid transporting conduits, air bubbles can be entrapped or particulates may become lodged in the networks. Nozzles of ordinary inkjet printheads are also subject to clogging, especially when used to eject a macromolecule-containing fluid at elevated temperatures, a situation commonly associated with such technologies. As a result, ordinary fluid dispensing technologies are prone to produce improperly sized or misdirected droplets.
[0004] A number of patents have described the use of focused acoustic radiation to dispense fluids such as inks and reagents. For example, U.S. Patent No. 4,308,547 to Lovelady et al. describes a liquid drop emitter that utilizes acoustic principles to eject droplets from a body of liquid onto a moving document to result in the formation of characters or barcodes thereon. A nozzleless inkjet printing apparatus is used such that controlled drops of ink are propelled by an acoustical force produced by a curved transducer at or below the surface of the ink. Similarly, U.S. Patent No. 6,666,541 to Ellson et al. describes a device for acoustically ejecting a plurality of fluid droplets toward discrete sites on a substrate surface for deposition thereon. The device includes an acoustic radiation generator for generating acoustic and a focusing means, e.g., a curved surface, for focusing acoustic radiation generated by the generator. In operation, the acoustic generator is acoustically coupled to the reservoir and activated to generate acoustic radiation. The focusing means then focuses the generated acoustic radiation at a point near a free fluid surface within the fluid contained in the reservoir. As a result, a fluid droplet is ejected from reservoir. U.S. Patent No. 6,596,239 to Williams describes technologies that employ focused acoustic technology as well.
[0005] Acoustic ejection provides a number of advantages over other fluid dispensing technologies. In contrast to inkjet devices, nozzleless fluid ejection devices are not subject to clogging and their associated disadvantages, e.g., misdirected fluid or improperly sized droplets. In contrast to inkjet printing devices, focused acoustic radiation may be used to effect nozzleless fluid ejection, and devices using focused acoustic radiation are not generally subject to clogging and the disadvantages associated therewith, e.g., misdirected fluid or improperly sized droplets. Furthermore, acoustic technology does not require the use of capillaries or involve invasive mechanical actions, for example, those associated with the introduction of a pipette tip into a reservoir of fluid. [0006] Acoustic radiation may also be used to assess the contents of one or more reservoirs. For example, the device described in U.S. Patent No. 6,666,541 to Ellson et al. may also be used to produce a detection acoustic wave that is transmitted to the fluid surface of the reservoir to become a reflected acoustic wave. Characteristics of the reflected acoustic radiation may then be analyzed in order to assess the spatial relationship between the acoustic radiation generator and the fluid surface. In addition, pool depth feedback technology using acoustic radiation is described in U.S. Patent No. 5,520,715 to Oeftering. Furthermore, U.S. Patent No. 6,596,239 to Williams describes acoustic ejection and pool depth detection technology. In some instances, detailed information relating to the contents of fluid in reservoirs may be obtained. For example, U.S. Patent Application Publication Nos. 20030101819 and 20030150257, each to Mutz et al., describe devices and methods for acoustically assessing the contents in a plurality of reservoirs.
[0007] As discussed above, when acoustic radiation is used to analyze the contents of a reservoir or to eject a fluid droplet therefrom, a generator for generating acoustic radiation is placed in acoustic coupling relationship with the reservoir. Although the generator may be placed within the reservoir to establish acoustic coupling, e.g., submerged in a fluid contained in the reservoir, submersion is undesirable when the acoustic generator is used to eject different fluids in rapid succession. Cleaning would be required to avoid contamination between the fluids. Thus, a preferred approach is to couple the generator to an exterior surface of the reservoir and to avoid placing the generator in the reservoir. As a result, the generator does not contact any fluid that the reservoir may contain.
[0008] For example, acoustic coupling may be achieved between an acoustic generator and a reservoir via an acoustic coupling medium. As described in U.S. Patent No. 6,666,541 to Ellson et al., such a coupling medium allows transmission of acoustic radiation therethrough and into the reservoir. Preferably, the acoustic coupling medium is an acoustically homogeneous fluid in conformal contact with both acoustic generator and the reservoir.
[0009] Since fluids used in pharmaceutical, biotechnological, and other scientific industries may be rare and/or expensive, techniques capable of handling small volumes of fluids provide readily apparent advantages over those requiring relatively larger volumes. Typically, fluids for use in combinatorial methods are provided as a collection or library of organic and or biological compounds. In many instances, well plates are used to store a large number of fluids for screening and/or processing. Well plates are typically of single piece construction and comprise a plurality of identical wells, wherein each well is adapted to contain a small volume of fluid. Such well plates are commercially available in standardized sizes and may contain, for example, 96, 384, 1536, or 3456 wells per well plate.
[0010] The ideal fluid-dispensing technique for pharmaceutical, biotechnological, medical (including clinical testing), and other industries provides for highly repeatable and accurate ejection of minute volumes of fluids directly from wells of a well plate. When used to prepare biomolecular arrays, the dispensing technique provides for deposition of droplets on a substrate surface, wherein droplet volume—and thus "spot" size on the substrate surface- can be carefully controlled. In order to ensure accurate placement of the droplets on a substrate surface, the droplets must take an appropriate trajectory from the wells of well plates towards the destination substrate.
[0011] The use of electric fields is well known in the printing arts to control the trajectory of ink droplets in a predetermined trajectory. For example, U.S. Patent 5,975,683 to Smith et al. describes a method and an apparatus that employ electrostatic acceleration to compensate for environmental factors that cause misdirection of ink droplets from an inkjet printhead. In addition, U.S. Patent No. 4,346,387 to Hertz describes a method and an apparatus for controlling the electrostatic charge on liquid droplets formed from a liquid stream emerging from a nozzle of an inkjet printhead.
[0012] Similarly, the use of electric fields is known in conjunction with focused acoustic radiation. For example, U.S. Patent Nos. 5,520,715 and 5,722,479, each to Oeftering, describe an apparatus for manufacturing a freestanding solid metal part through acoustic ejection of charged molten metal droplets. The apparatus employs electric fields to direct the charged droplets to predetermined points on a target where the droplets solidify as a result of cooling. Similarly, U.S. Patent Application Publication Nos. 20020109084 and 20020125424, each to Ellson et al., describe the use of focused acoustic radiation to introduce droplets of fluids into ionization chambers such as those associated with mass spectrometers. Moreover, U.S. Patent Nos. 6,079,814 and 6,367,909, each to Lean et al., describe printing methods and apparatuses that employ electric fields to reduce drop placement errors. Typically, an aperture plate is used to charge a free surface of a fluid in a reservoir. Then, focused acoustic radiation is applied to a point near the fluid surface so as to eject a charged droplet therefrom and through the aperture of the plate. Additional electric fields may be employed to direct the charged droplet so that it follows a predetermined trajectory. Optionally, an electric field may also serve to tack a recording medium in position to receive the ink droplet.
[0013] Although it is sometimes a straightforward matter to use electric fields to control the size and trajectory of droplet ejected from a single reservoir, it is quite difficult to achieve such control in high-throughput applications. For example, when acoustic ejection is employed to transfer fluids from a 96- well source plate to a 384-well target plate, the relative motion between the plates makes it difficult to maintain the presence of a consistent charge within each well over time. In addition, it has been discovered that wells of commercially available well plates, particularly those made from plastic materials such as polypropylene, polystyrene, or cyclic olefins, are often prone to accumulate uncontrolled electrostatic charge. Uncontrolled electrostatic charge tends to alter the volume and/or trajectory of droplets dispensed from well plates. This alteration in droplet volume and/or trajectory particularly pronounced for devices constructed to dispense droplets at a relatively low velocity.
[0014] Thus, there is a need to reduce the accumulation of uncontrolled electrostatic charge associated with droplet-dispensing devices, in order to control the volume and/or trajectory of a droplet dispensed from a reservoir of such a device. Since droplets ejected using focused acoustic radiation tends to exhibit a lower velocity than droplets ejected from ordinary inkjet technologies such as thermal ejection, the need is particularly great for ejection devices that use focused acoustic radiation such as those employed to effect highspeed monitoring and/or ejection of fluid in a plurality of reservoirs.
DISCLOSURE OF THE INVENTION [0015] Accordingly, it is an object of the present invention to provide devices and methods that overcome the above-mentioned disadvantages of the prior art. [0016] In one embodiment, the invention provides a device comprised of a reservoir adapted to contain a fluid and a dispenser for dispensing a fluid droplet from the reservoir. A means is employed for reducing uncontrolled electrostatic charge on the reservoir when the reservoir is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom. The means for reducing uncontrolled electrostatic charge is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory. Often grounding is used to reduce or eliminate uncontrolled electrostatic charge.
[0017] In another embodiment, the invention provides a similar device that further comprises a substrate positioned to receive the dispensed droplet. When the substrate is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of the dispensed droplet, a means for reducing uncontrolled electrostatic charge is provided that is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
[0018] Typically, the dispenser is comprised of an acoustic ejector. In some instances, the acoustic ejector may comprise an acoustic radiation generator for generating acoustic radiation and a focusing means for focusing the acoustic radiation generated. In such cases, the invention also provides a means for positioning the ejector in acoustic coupling relationship to the reservoir. Typically, the reservoir, the substrate, and any other component of the device prone to accumulate uncontrolled electrostatic charge have an electrical resistivity of no more than about 1011 ohm-cm, have a surface electrical resistivity of no more than about 1012 ohm/sq, or both. This may be achieved by using a material that is at least partially nonmetallic or polymeric.
[0019] In a further embodiment, the invention provides a method for dispensing a droplet from a reservoir containing a fluid. The method involves reducing uncontrolled electrostatic charge on the reservoir when the reservoir is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom. As a result, uncontrolled electrostatic charge is reduced to a level effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory. [0020] In yet another embodiment, the invention provides a method for dispensing a droplet from a reservoir containing a fluid onto a substrate. The method involves reducing uncontrolled electrostatic charge on the reservoir and/or the substrate when the reservoir and/or substrate are prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of the dispensed droplet. Uncontrolled electrostatic charge is reduced to a level effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
[0021] For any of the inventive methods, focused acoustic radiation may be applied in a manner effective to eject a droplet of fluid from the reservoir.
BRΓEF DESCRΓPTIQN OF THE DRAWINGS
[0022] The invention is described in detail below with reference to the following drawings, wherein like reference numerals indicate a corresponding structure throughout the several views.
[0023] FIG. 1A and FIG. IB, collectively referred to as FIG. 1, schematically illustrate in simplified cross-sectional view the operation of a focused acoustic ejection device in the preparation of a plurality of features on a substrate surface. FIG. 1A shows the acoustic ejector acoustically coupled to a first reservoir and having been activated in order to eject a first droplet of fluid from within the reservoir toward a particular site on a substrate surface. FIG. IB shows the acoustic ejector acoustically coupled to a second reservoir and having been activated to eject a second droplet of fluid from within the second reservoir.
[0024] FIG. 2 illustrates in cross-sectional schematic view the ejection of droplets of fluid from a volume of fluid on a substrate surface into an inlet opening disposed on a terminus of a capillary.
[0025] FIG. 3A, FIG. 3B, FIG. 3C and FIG. 3D, collectively referred to as FIG. 3, schematically illustrate in simplified cross-sectional view a known device that employs an acoustic coupling fluid and the disadvantages associated therewith. As depicted, the device comprises first and second reservoirs, a combined acoustic analyzer and ejector unit, and an ejector positioning means. FIG. 3A shows the acoustic unit acoustically coupled to the first reservoir so that the unit is activated to determine the position of the free fluid surface within the first reservoir. FIG. 3B depicts the repositioning of the acoustic unit toward the reservoir and the activation acoustic unit in order to eject a droplet of fluid from within the first reservoir toward a site on a substrate surface to form an array. FIG. 3C shows the acoustic unit acoustically coupled to the second reservoir so that the unit is activated to determine the position of the free fluid surface within the second reservoir. FIG. 3D depicts the repositioning of the acoustic unit away from the reservoir and the activation acoustic unit in order to eject a droplet of fluid from within the second reservoir toward a site on a substrate surface.
[0026] FIG. 4 schematically illustrate in simplified cross-sectional view a device that includes a nozzle located within a collector.
[0027] FIG. 5 schematically illustrates in simplified cross-sectional view an acoustic device having a dispenser that employs a stationary opposing piston design.
[0028] FIG. 6 schematically illustrates in simplified cross-sectional view an acoustic device similar to that of FIG. 3 except that the acoustic ejector and the positioning means are sealed and in a container filled completely with the acoustic coupling fluid
DETAILED DESCRD?TIQN OF THE INVENTION
[0029] Before describing the present invention in detail, it is to be understood that this invention is not limited to specific fluids, biomolecules, or device structures, as such may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.
[0030] It must be noted that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include both singular and plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a reservoir" includes a plurality of reservoirs as well as a single reservoir, reference to "a droplet" includes a plurality of droplets as well as single droplet, and the like. [0031] In describing and claiming the present invention, the following terminology will be used in accordance with the definitions set out below.
[0032] The terms "acoustic coupling" and "acoustically coupled" as used herein refer to a state wherein an object is placed in direct or indirect contact with another object so as to allow acoustic radiation to be transferred between the objects without substantial loss of acoustic energy. When two items are indirectly acoustically coupled, an "acoustic coupling medium" is needed to provide an intermediary through which acoustic radiation may be transmitted. Thus, an ejector may be acoustically coupled to a fluid, e.g., by immersing the ejector in the fluid or by interposing an acoustic coupling medium between the ejector and the fluid, in order to transfer acoustic radiation generated by the ejector through the acoustic coupling medium and into the fluid. In some instances, the acoustic coupling medium may include or be formed entirely of a solid material.
[0033] The term "array" as used herein refers to a two-dimensional arrangement of features, such as an arrangement of reservoirs (e.g., wells in a well plate) or an arrangement of different moieties, including ionic, metallic, or covalent crystalline, e.g., molecular crystalline, composite, ceramic, vitreous, amorphous, fluidic, or molecular materials on a substrate surface (as in an ohgonucleotide or peptidic array). Arrays are generally comprised of regular, ordered features, as in, for example, a rectilinear grid, parallel stripes, spirals, and the like, but non-ordered arrays may be advantageously used as well. In particular, the term "rectilinear array" as used herein refers to an array that has rows and columns of features wherein the rows and columns typically, but not necessarily, intersect each other at a ninety- degree angle. An array is distinguished from the more general term "pattern" in that patterns do not necessarily contain regular and ordered features. Arrays typically but do not necessarily comprise at least about 4 to about 10,000,000 features, generally in the range of about 4 to about 1,000,000 features.
[0034] The term "attenuation" is used herein in its ordinary sense and refers to the decrease in intensity of a wave due to scattering and/or absorption of energy. Typically, attenuation occurs with little or no distortion but does not include intensity reduction due to geometric spreading. Thus, the term "attenuation coefficient" refers to the rate of diminution of wave intensity with respect to distance along a transmission path. [0035] The terms "biomolecule" and "biological molecule" are used interchangeably herein to refer to any organic molecule that is, was, or can be a part of a living organism, regardless of whether the molecule is naturally occurring, recombinantly produced, or chemically synthesized in whole or in part. The terms encompass, for example, nucleotides, amino acids, and monosaccharides, as well as oligomeric and polymeric species, such as oligonucleotides and polynucleotides; peptidic molecules, such as oligopeptides, polypeptides, and proteins; saccharides, such as disaccharides, oligosaccharides, polysaccharides, and mucopolysaccharides or peptidoglycans (peptido-polysaccharides); and the like. The terms also encompass ribosomes, enzyme cofactors, pharmacologically active agents, and the like. Additional information relating to the term "biomolecule" can be found in U.S. Patent No. 6,666,541 to Ellson et al.
[0036] The term "capillary" is used herein to refer to a conduit having a bore of small dimension. Typically, capillaries for electrophoresis that are free standing tubes have an inner diameter in the range of about 50 to about 250 μm. Capillaries with extremely small bores integrated to other devices, such as openings for loading microchannels of microfluidic devices, can be as small as 1 μm, but in general these capillary openings are in the range of about 10 to about 100 μm. In the context of delivery to a mass analyzer in electrospray-type mass spectrometry, the inner diameter of capillaries may range from about 0.1 to about 3 mm and preferably from about 0.5 to about 1 mm. In some instances, a capillary can represent a portion of a microfluidic device. In such instances, the capillary may be an integral or affixed (permanently or detachably) portion of the microfluidic device.
[0037] The term "fluid" as used herein refers to matter that is nonsolid, or at least partially gaseous and/or liquid, but not entirely gaseous. A fluid may contain a solid that is minimally, partially, or fully solvated, dispersed, or suspended. Examples of fluids include, without limitation, aqueous liquids (including waterier se and salt water) and nonaqueous liquids such as organic solvents and the like. As used herein, the term "fluid" is not synonymous with the term "ink" in that an ink must contain a colorant and may not be gaseous.
[0038] The terms "focusing means" and "acoustic focusing means" refer to a means for causing acoustic waves to converge at a focal point, either by a device separate from the acoustic energy source that acts like an optical lens, or by the spatial arrangement of acoustic energy sources to effect convergence of acoustic energy at a focal point by constructive and destructive interference. A focusing means may be as simple as a solid member having a curved surface, or it may include complex structures such as those found in Fresnel lenses, which employ diffraction in order to direct acoustic radiation. Suitable focusing means also include phased array methods as are known in the art and described, for example, in U.S. Patent No. 5,798,779 to Nakayasu et al. and by Amemiya et al. (1997) Proceedings of the 1997 IS&TNIP13 International Conference on Digital Printing Technologies, pp. 698-702. Additional information regarding acoustic focusing is provided in U.S. Patent Application Publication No. 20020125424 to Ellson et al.
[0039] The terms "library" and "combinatorial library" are used interchangeably herein to refer to a plurality of chemical or biological moieties arranged in a pattern or an array such that the moieties are individually addressable. In some instances, the plurality of chemical or biological moieties is present on the surface of a substrate, and in other instances the plurality of moieties represents the contents of a plurality of reservoirs. Preferably, but not necessarily, each moiety is different from each of the other moieties. The moieties may be, for example, peptidic molecules and/or oligonucleotides.
[0040] The "limiting dimension" of an opening refers herein to the theoretical maximum diameter of a sphere that can pass through an opening without deformation. For example, the limiting dimension of a circular opening is the diameter of the opening. As another example, the limiting dimension of a rectangular opening is the length of the shorter side of the rectangular opening. The opening may be present on any solid body including, but not limited to, sample vessels, substrates, capillaries, microfluidic devices, and ionization chambers. Depending on the purpose of the opening, the opening may represent an inlet and/or an outlet.
[0041] The term "moiety" refers to any particular composition of matter, e.g., a molecular fragment, an intact molecule (including a monomeric molecule, an oligomeric molecule, or a polymer), or a mixture of materials (for example, an alloy or a laminate).
[0042] The term "near," as used herein, refers to the distance from the focal point of the focused acoustic radiation to the surface of the fluid from which a droplet is to be ejected, and indicates that the distance should be such that the focused acoustic radiation directed into the fluid results in droplet ejection from the fluid surface; one of ordinary skill in the art will be able to select an appropriate distance for any given fluid using straightforward and routine experimentation. Generally, however, a suitable distance between the focal point of the acoustic radiation and the fluid surface is in the range of about 1 to about 15 times the wavelength of the acoustic radiation in the fluid, more typically in the range of about 1 to about 10 times that wavelength, preferably in the range of about 1 to about 5 times that wavelength.
[0043] "Optional" or "optionally" means that the subsequently described circumstance may or may not occur, so that the description includes instances where the circumstance occurs and instances where it does not.
[0044] The term "radiation" is used in its ordinary sense and refers to emission and propagation of energy in the form of a waveform disturbance traveling through a medium such that energy is transferred from one particle of the medium to another, generally without causing any permanent displacement of the medium itself. Thus, radiation may refer, for example, to electromagnetic waveforms as well as acoustic vibrations.
[0045] Accordingly, the terms "acoustic radiation" and "acoustic energy" are used interchangeably herein and refer to the emission and propagation of energy in the form of sound waves. As with other waveforms, acoustic radiation may be focused using a focusing means, as discussed below. Although acoustic radiation may have a single frequency and associated wavelength, acoustic radiation may take a form, e.g. a "linear chirp," that includes a plurality of frequencies. Thus, the term "characteristic wavelength" is used to describe the mean wavelength of acoustic radiation having a plurality of frequencies.
[0046] The term "reservoir" as used herein refers to a receptacle or chamber for containing a fluid. In some instances, a fluid contained in a reservoir necessarily will have a free surface, e.g., a surface that allows acoustic radiation to be reflected therefrom or a surface from which a droplet may be acoustically ejected. A reservoir may also be a locus on a substrate surface within which a fluid is constrained.
[0047] The term "substrate" as used herein refers to any material having a surface onto which one or more fluids may be deposited. The substrate may be constructed in any of a number of forms including, for example, wafers, slides, well plates, or membranes. In addition, the substrate may be porous or nonporous as required for deposition of a particular fluid. Suitable substrate materials include, but are not limited to, supports that are typically used for solid phase chemical synthesis, such as polymeric materials (e.g., polystyrene, polyvinyl acetate, polyvinyl chloride, polyvinyl pyrrolidone, polyacrylonitrile, polyacrylamide, polymethyl methacrylate, polytetrafluoroethylene, polyethylene, polypropylene, polyvinylidene fluoride, polycarbonate, and divinylbenzene styrene-based polymers), agarose (e.g., Sepharose®), dextran (e.g., Sephadex®), cellulosic polymers and other polysaccharides, silica and silica-based materials, glass (particularly controlled pore glass, or "CPG") and functionalized glasses, ceramics, and such substrates treated with surface coatings, e.g., with microporous polymers (particularly cellulosic polymers such as nitrocellulose), microporous metallic compounds (particularly microporous aluminum), antibody-binding proteins (available from Pierce Chemical Co., Rockford IL), bisphenol A polycarbonate, or the like. Additional information relating to the term "substrate" can be found in U.S. Patent No. 6,666,541 to Ellson et al.
[0048] The term "substantially" as in, for example, the phrase "substantially deviate from a predetermined volume," refers to a volume that does not deviate by more than about 25%, preferably 10%, more preferably 5%, and most preferably at most 2%, from the predetermined volume. Other uses of the term "substantially" involve an analogous definition.
[0049] The term "sample vessel" as used herein refers to any hollow or concave receptacle having a structure that allows for sample processing and/or analysis. Thus, a sample vessel has an inlet opening through which sample may be introduced and an optional, but preferred, outlet opening through which processed or analyzed sample may exit.
[0050] In general, the invention relates to devices and methods for dispensing a fluid droplet of a predetermined volume and/or predetermined trajectory from a reservoir adapted to contain a fluid. The invention derives from the observation that fluid dispensing devices or components thereof sometimes accumulate uncontrolled electrostatic charge such that droplets dispensed therefrom exhibit a volume and/or trajectory that substantially deviate from the predetermined volume and/or predetermined trajectory. This is particularly problematic when the device is adapted to dispense droplets containing a minute volume of fluid. Often, the reservoir itself is prone to accumulate such uncontrolled electrostatic charge. Thus, the invention provides for the reduction of such uncontrolled electrostatic charge in a manner effective to ensure that the volume and/or trajectory of the dispensed droplet conform to the predetermined volume and/or trajectory. In particular, the invention is particularly suited for applications that require the efficient transport and/or deposition of small quantities of fluid.
[0051] Among the various routes for an item to accumulate electrostatic charge is the triboelectric effect, by which an item will typically accumulate uncontrolled electrostatic charge through friction, pressure, and separation. The magnitude of the static charge is typically determined by material composition, applied forces, separation rate, and dissipative forces. Generally, the ability of a material to surrender or gain electrons is a function of the conductivity of the material. The tendency of a material to accumulate uncontrolled electrostatic charge is inversely correlated to the surface and/or volume conductivity of the material. Accordingly, the invention is particularly suited for use in devices comprised of components that exhibit a low electrical conductivity or high electrical resistivity. Typically, the invention will be useful to reduce uncontrolled electrostatic charge in items having a volume electrical resistivity of at least 10 ohm-cm and/or a surface electrical resistivity of at least 1014 ohm/sq. As the usefulness of the invention increases with the electrical resistance of the item requiring reduction in controlled electrostatic charge, one skilled in the art will recognize that the invention will be particularly useful to discharge items having a volume electrical resistivity of at least 1015 or 1016 ohm-cm and/or a surface electrical resistivity of at least 1016 or 1017 ohm/sq.
[0052] The invention may be employed with any type of fluid dispenser that serves to dispense one or more droplets of fluid from a reservoir. Any fluid droplet dispensing techniques known in the art may be used in conjunction with the present invention. For example, the invention may be used with dispensers such as inkjet printheads (both thermal and piezoelectric), pipettes, capillaries, syringes, displacement pumps, rotary pumps, peristaltic pumps, vacuum devices, flexible or rigid tubing, valves, manifolds, pressurized gas canisters, and combinations thereof. While nonacoustic techniques may be used to dispense fluid from the reservoir, the invention is particularly suited for use with nozzleless acoustic ejection techniques that employ focused acoustic radiation generated by acoustic ejectors, such as those described in U.S. Patent No. 6,666,541 to Ellson et al. This publication sets forth that an ejector may be acoustically coupled to a reservoir containing a fluid in order to eject a droplet therefrom. In some instances, the reservoir may be a well of a well plate. Since this device configuration allows droplets to be ejected from near the base of a well, uncontrolled electrostatic charge anywhere in the well, e.g., the base or sidewalls, may have a strong effect influence on the volume and or trajectory of such droplets. Since current inkjet systems do not typically exhibit such a configuration, devices having such a configuration, e.g., devices that employ focused acoustic radiation, may benefit more from the invention than ordinary inkjet technologies.
[0053] Since acoustic ejection provides a number of advantages over other fluid dispensing technologies, one embodiment of the invention provides a device for acoustically ejecting a droplet of fluid from a reservoir. The device is comprised of a reservoir adapted to contain a fluid, an ejector for ejecting a droplet from the reservoir, and a means for positioning the ejector in acoustic coupling relationship to the reservoir. The ejector comprises an acoustic radiation generator for generating acoustic radiation and a focusing means for focusing the acoustic radiation generated by the generator. As described in U.S. Patent No. 6,666,541 to Ellson et al., the acoustic radiation is focused at a focal point within and sufficiently near the fluid surface in the reservoir to result in the ejection of droplets therefrom. Furthermore, a means is provided for reducing any uncontrolled electrostatic charge on the device or a portion thereof that alters the volume and or trajectory of a droplet ejected from the reservoir. As a result, the volume and/or trajectory of the ejected droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
[0054] The device may be constructed to include the reservoir as an integrated or permanently attached component of the device. However, to provide modularity and interchangeability of components, it is preferred that the device be constructed with a removable reservoir. Optionally, a plurality of reservoirs many be provided. Generally, the reservoirs are arranged in a pattern or an array to provide each reservoir with individual systematic addressability. In addition, while each of the reservoirs may be provided as a discrete or stand-alone item, in circumstances that require a large number of reservoirs, it is preferred that the reservoirs be attached to each other or represent integrated portions of a single reservoir unit. For example, the reservoirs may represent individual wells in a well plate. [0055] Many well plates suitable for use with the device are commercially available and may contain, for example, 96, 384, 1536, or 3456 wells per well plate, having a full skirt, half skirt, or no skirt. The wells of such well plates typically form rectilinear arrays. Manufacturers of suitable well plates for use in the employed device include Corning, Inc. (Corning, New York) and Greiner America, Inc. (Lake Mary, Florida). However, the availability of such commercially available well plates does not preclude the manufacture and use of custom-made well plates containing at least about 10,000 wells, or as many as 100,000 to 500,000 wells, or more. The wells of such custom-made well plates may form rectilinear or other types of arrays. As well plates have become commonly used laboratory items, the Society for Biomolecular Screening (Danbury, Connecticut) has formed the Microplate Standards Development Committee to recommend and maintain standards to facilitate the automated processing of small volume well plates on behalf of and for acceptance by the American National Standards Institute.
[0056] Furthermore, the material used in the construction of reservoirs must be compatible with the fluids contained therein. Thus, if it is intended that the reservoirs or wells contain an organic solvent such as acetonitrile, polymers that dissolve or swell in acetonitrile would be unsuitable for use in forming the reservoirs or well plates. Similarly, reservoirs or wells intended to contain DMSO must be compatible with DMSO. For water- based fluids, a number of materials are suitable for the construction of reservoirs and include, but are not limited to, ceramics such as silicon oxide and aluminum oxide, metals such as stainless steel and platinum, and polymers such as polyester and polytetrafluoroethylene. For fluids that are photosensitive, the reservoirs may be constructed from an optically opaque material that has sufficient acoustic transparency for substantially unimpaired functioning of the device. Thus, the reservoir may be adapted to contain any type of fluid, metallic or nonmetallic, organic or inorganic.
[0057] It should be noted that from a manufacturing perspective, polymeric materials are particularly suited for use in forming reservoirs for use with the invention, e.g., well plates that conform to industrial standards. Such materials typically exhibit the appropriate mechanical, acoustical, and chemical properties suited for use with the invention. For example, well plates may be formed from polymeric material selected from the group consisting of polyethylenes, polypropylenes, polybutylenes, polystyrenes, cyclic olefins, combinations thereof, and copolymers of any of the foregoing. Such polymers are generally inert to aqueous solutions and can be easily formed through casting, injection molding, extrusion, and other well-established processing techniques. However, such polymers are noted for their high volume and surface resistivity, e.g., at least 1013 ohm-cm and at least 1014 ohm/sq, respectively. Thus, the invention also relates to reservoirs and well plates that exhibit a resistivity wherein the reservoir, the optional substrate, or both are comprised of a material that is at least partially polymeric and either has an electrical resistivity of no more than about 10 ohm-cm, has a surface electrical resistivity of no more than about 10 ohm/sq, or both.
[0058] While most polymeric materials are insulators, conductive polymers are known in the art. For example, polythiophenes are a well-known class of conductive polymer and generally exhibit greater chemical stability than polyacetylene derivatives. Conductive polymer materials are extremely economical to produce and have been used commercially in the semiconductor field as containers for electrostatically sensitive materials. Relatively stable polythiophene derivatives include polyisothianapthene (PITN) and poly-3,4,ethylene dioxythiophene (PEDT), and a variety of related materials such as doped polypropylenes, are commercially available from RTP Company (Winona, Minnesota).
[0059] In some instances, an electrically conductive layer may be used to increase the conductivity of a reservoir. Such a layer may be provided as a surface coating or incorporated within a reservoir to increase the reservoir's conductivity. For example, any part of an ordinary plastic well plate comprising an array of 96 substantially identical wells prone to accumulate uncontrolled electrostatic charge may be coated with a metallic coating. For example, metals such as aluminum, gold, silver, copper, platinum, palladium, or nickel may be selectively deposited on the upper, lower, interior, and/or exterior surface of an ordinary commercially available well plate. Similarly, plating technologies may be used to increase the thickness of the metallic coating. Furthermore, nonmetallic coatings may be used as well. For example, known conductive ceramic coating materials include indium tin oxide and titanium nitride. In addition, various forms of carbon, e.g., carbon fibers, graphite, or acetylene black, may be applied as a surface coating on the reservoir.
[0060] In addition, or in the alternative, a polymeric reservoir may contain an electrically conductive filler. Any of the materials suitable for forming the electrically conductive layer as discussed above may be used as a filler material. For example, carbon- filled plastics are well known in the art for electrostatic dissipation. Such carbon-filled plastics may be obtained from Minnesota Mining & Manufacturing Company Corporation (St. Paul, Minnesota) under the trademark Nelostat®. Such reservoirs may be formed using ordinary polymer processing techniques.
[0061] When a plurality of reservoirs is employed, the acoustic radiation generator may have to be aligned with each reservoir during operation, discussed infra. In order to reduce the amount of movement and time needed to align the generator successively with each reservoir, it is preferable that the center of each reservoir be located not more than about 1 centimeter, more preferably not more than about 1.5 millimeters, still more preferably not more than about 1 millimeter and optimally not more than about 0.5 millimeter, from a neighboring reservoir center. These dimensions tend to limit the size of the reservoirs to a maximum volume. The reservoirs are constructed to contain typically no more than about 1 mL, preferably no more than about 100 μL, more preferably no more than about 10 μL, still more preferably no more than about 1 μL, and optimally no more than about 1 nL, of fluid. The reservoirs may be either completely or partially filled with fluid. For example, fluid may occupy a volume of about 10 pL to about 100 nL.
[0062] When an array of reservoirs is provided, each reservoir may be individually, efficiently, and systematically addressed. Although any type of array may be employed, arrays comprised of parallel rows of evenly spaced reservoirs are preferred. Typically, though not necessarily, each row contains the same number of reservoirs. Optimally, rectilinear arrays comprising X rows and Y columns of reservoirs are employed with the invention, wherein X and Y are each at least 2. In some instances, X may be greater than, equal to, or less than Y. In addition, nonrectilinear arrays as well as other geometries may be employed. For example, hexagonal, spiral, or other types of arrays may be used. In some instances, the invention may be employed with irregular patterns of reservoirs, e.g., droplets randomly located on a flat substrate surface such as those associated with a CD-ROM format. In addition, the invention may be used with reservoirs associated with microfluidic devices.
[0063] Moreover, the invention may be used to dispense fluids of virtually any type and amount desired. The fluid may be aqueous and/or nonaqueous. Examples of fluids include, but are not limited to, aqueous fluids including waterier se and water-solvated ionic and non-ionic solutions; organic solvents; lipidic liquids; suspensions of immiscible fluids; and suspensions or slurries of solids in liquids. Because the invention is readily adapted for use with high temperatures, fluids such as liquid metals, ceramic materials, and glasses may be used, as described in U.S. Patent Application Publication No. 20020140118. In some instances, the reservoir may contain a biomolecule, nucleotidic, peptidic, or otherwise. In addition, the invention may be used in conjunction with dispensers for dispensing droplets of immiscible fluids, as described in U.S. Patent Application Publication Nos. 2002037375 and 20020155231, or to dispense droplets containing pharmaceutical agents, as discussed in U.S. Patent Application Publication No. 20020142049 and U.S. Patent Application Publication No. 20030012892 to et al.
[0064] Any of a variety of focusing means may be employed to focus acoustic radiation so as to eject droplets from a reservoir. For example, one or more curved surfaces may be used to direct acoustic radiation to a focal point near a fluid surface. One such technique is described in U.S. Patent No. 4,308,547 to Lovelady et al. Focusing means with a curved surface have been incorporated into the construction of commercially available acoustic transducers such as those manufactured by Panametrics Inc. (Waltham, Massachusetts). In addition, Fresnel lenses are known in the art for directing acoustic energy at a predetermined focal distance from an object plane. See, e.g., U.S. Patent No. 5,041,849 to Quate et al. Fresnel lenses may have a radial phase profile that diffracts a substantial portion of acoustic energy into a predetermined diffraction order at diffraction angles that vary radially with respect to the lens. The diffraction angles should be selected to focus the acoustic energy within the diffraction order on a desired object plane. It should be noted that acoustic focusing means exhibiting a variety of F-numbers may be employed with the invention. As discussed in U.S. Patent No. 6,416,164 to Stearns et al., however, low F- number focusing places restrictions on the reservoir and fluid level geometry and provides relatively limited depth of focus, increasing the sensitivity to the fluid level in the reservoir. Thus, the focusing means suitable for use with the invention typically exhibits an F-number of at least about 1. Preferably, the focusing means exhibits an F-number of at least about 2.
[0065] There are a number of ways to acoustically couple the ejector to a reservoir and thus to the fluid therein. One such approach is through direct contact, as is described, for example, in U.S. Patent No. 4,308,547 to Lovelady et al., wherein a focusing means constructed from a hemispherical crystal having segmented electrodes is submerged in a liquid to be ejected. The aforementioned patent further discloses that the focusing means may be positioned at or below the surface of the liquid. However, this approach for acoustically coupling the focusing means to a fluid is undesirable when the ejector is used to eject different fluids in a plurality of containers or reservoirs, as repeated cleaning of the focusing means would be required in order to avoid cross-contamination. The cleaning process would necessarily lengthen the transition time between each droplet ejection event. In addition, in such a method, fluid would adhere to the ejector as it is removed from each container, wasting material that may be costly or rare.
[0066] Thus, a preferred approach is to acoustically couple the ejector to the reservoir without contacting any portion of the ejector, e.g., the focusing means, with the fluids to be . ejected. When a plurality of reservoirs is employed, a positioning means is provided for positioning the ejector in controlled and repeatable acoustic coupling with each of the fluids in the reservoirs to eject droplets therefrom without submerging the ejector therein. This typically involves direct or indirect contact between the ejector and the external surface of each reservoir. When direct contact is used in order to acoustically couple the ejector to each reservoir, it is preferred that the direct contact be wholly conformal to ensure efficient acoustic energy transfer. That is, the ejector and the reservoir should have corresponding surfaces adapted for mating contact. Thus, if acoustic coupling is achieved between the ejector and reservoir through the focusing means, it is desirable for the reservoir to have an outside surface that corresponds to the surface profile of the focusing means. Without conformal contact, efficiency and accuracy of acoustic energy transfer may be compromised. In addition, since many focusing means have a curved surface, the direct contact approach may necessitate the use of reservoirs having a specially formed inverse surface.
[0067] When an ejector is placed in indirect contact with a reservoir, an acoustic coupling medium may be interposed between the reservoir and ejector. Typically, the acoustic coupling medium is a fluid. In addition, the acoustic coupling medium is preferably an acoustically homogeneous material that is substantially free of material having different acoustic properties than the fluid medium itself. Furthermore, it is preferred that the acoustic coupling medium be comprised of a material having acoustic properties that facilitate the transmission of acoustic radiation without significant attenuation in acoustic pressure and intensity. Also, the acoustic impedance of the coupling medium should facilitate the transfer of energy from the coupling medium into the reservoir. An aqueous fluid, such as waterier se, may be employed as an acoustic coupling medium. Ionic additives, e.g., salts, may sometimes be added to the coupling medium to increase the conductivity of the coupling medium, thereby facilitating discharge of any charge accumulated by the reservoir.
[0068] A single ejector is preferred, although the inventive device may include a plurality of ejectors. When a single ejector is employed, the means for positioning the ejector may be adapted to provide relative motion between the ejector and reservoirs. The positioning means should allow for the ejector to move from one reservoir to another quickly and in a controlled manner, thereby allowing fast and controlled scanning of the reservoirs to effect droplet ejection therefrom. Thus, various means for positioning the ejector in acoustic coupling relationship to the reservoir are generally known in the art and may involve, e.g., devices that provide movement having one, two, three, four, five, six, or more degrees of freedom. Accordingly, when rows of reservoirs are provided, relative motion between the acoustic radiation generator and the reservoirs may result in displacement of the acoustic radiation generator in a direction along the rows. Similarly, when a rectilinear array of reservoirs is provided, the ejector may be movable in a row- wise direction and/or in a direction perpendicular to both the rows and columns.
[0069] In addition, the rate at which fluid droplets can be delivered is related to the efficiency of fluid delivery. Current positioning technology allows for the ejector positioning means to move from one reservoir to another quickly and in a controlled manner, thereby allowing fast and controlled ejection of different fluid samples. That is, current commercially available technology allows the ejector to be moved from one reservoir to another, with repeatable and controlled acoustic coupling at each reservoir, in less than about 0.1 second for high performance positioning means and in less than about 1 second for ordinary positioning means. A custom designed system will allow the ejector to be moved from one reservoir to another with repeatable and controlled acoustic coupling in less than about 0.001 second.
[0070] The invention also enables rapid ejection of droplets from one or more reservoirs, e.g., at a rate of at least about 1,000,000 droplets per minute from the same reservoir, and at a rate of at least about 100,000 drops per minute from different reservoirs, assuming that the droplet size does not exceed about 10 μm in diameter. One of ordinary skill in the art will recognize that the droplet generation rate is a function of drop size, viscosity, surface tension, and other fluid properties. In general, the droplet generation rate increases with decreasing droplet diameter, and 1,000,000 droplets per minute is achievable for most aqueous fluid drops under about 10 μm in diameter.
[0071] The invention may be used in any context where precise placement of a fluid droplet is desirable or necessary. In particular, the invention may be employed to improve accuracy and precision associated with nozzleless acoustic ejection. For example, it is described in U.S. Patent No. 6,666,541 to Ellson et al. that acoustic ejection technology may be used to form biomolecular arrays. Similarly, acoustic ejection technology may be employed to format a plurality of fluids, e.g., to transfer fluids from odd-sized bulk containers to wells of a standardized well plate or to transfer fluids from one well plate to another. Furthermore, as described in U.S. Patent Application Publication Nos. 20020109084 and 20020125424, each to Ellson et al., focused acoustic radiation may serve to eject a droplet of fluid from a reservoir into any sample vessel for processing and/or analyzing a sample molecule, e.g., into a sample introduction interface of a mass spectrometer, an inlet opening that provides access to the interior region of a capillary, or an inlet port of a microfluidic device. Similarly, the invention may be used to eject droplets of analysis-enhancing fluid on a sample surface in order to prepare the sample for analysis, e.g., for MALDI or SELDI-type analysis.
[0072] As discussed above, uncontrolled electrostatic charge may be accumulated by a substrate onto which droplets are dispensed. Such charge may also have a detrimental influence on the trajectory and/or volume of the dispensed droplets. Thus, construction considerations for such substrates are similar to those associated with reservoirs, as discussed above. For example, the substrate may exhibit a relatively high electrical conductivity for ease in grounding. Similarly, the materials and techniques suitable for use in forming the reservoir may also be used with the substrate. In some instances, a single means for reducing uncontrolled charge may be used for both the reservoir and substrate.
[0073] In order to prepare an array on a substrate surface, the substrate must be placed in droplet-receiving relationship to a reservoir. Thus, the invention may also employ a positioning means for positioning the substrate. With respect to the substrate positioning means and the ejector positioning means, it is important to keep in mind that there are two basic kinds of motion: pulse and continuous. For the ejector positioning means, pulse motion involves the discrete steps of moving an ejector into position, emitting acoustic energy, and moving the ejector to the next position; again, using a high performance positioning means with such a method allows repeatable and controlled acoustic coupling at each reservoir in less than 0.1 second. A continuous motion design, on the other hand, moves the ejector and the reservoirs continuously, although not necessarily at the same speed, and provides for ejection during movement. Since the pulse width is very short, this type of process enables over 10 Hz reservoir transitions, and even over 1000 Hz reservoir transitions. In any case, relative motion between the reservoirs and the acoustic generator can be achieved by moving the reservoirs while holding the generator still, by moving the reservoirs while holding the generator still, or by moving the generator and the reservoirs at different velocities. Similar engineering considerations are applicable to the substrate positioning means.
[0074] From the above, it is evident that the relative positions and spatial orientations of the various components may be altered depending on the particular desired task at hand. In such a case, the various components of the device may require individual control or synchronization to direct droplets onto designated sites on a substrate surface. For example, the ejector positioning means may be adapted to eject droplets from each reservoir in a predetermined sequence associated with an array of designated sites on the substrate surface. Any positioning means of the present invention may be constructed from, e.g., levers, pulleys, gears, a combination thereof, or other mechanical means known to one of ordinary skill in the art.
[0075] In some instances, acoustic energy may be used to assess the contents of a plurality of reservoirs. Such acoustic assessment technologies are described U.S. Patent Application Publication Nos. 20030101819 and 20030150257, each to Mutz et al. For example, the acoustic radiation generator may be used in combination with an analyzer for analyzing a characteristic of acoustic radiation generated by the generator and transmitted through the reservoir. By placing the analyzer in radiation receiving relationship to the acoustic radiation generator, the acoustic radiation having interacted with the contents of the reservoir may be analyzed. Such acoustic assessment of the contents of one or more reservoirs may enhance the accuracy and precision in dispensing fluids therefrom. [0076] A means for reducing uncontrolled electrostatic charge is employed so that any dispensed droplet exhibits a volume and/or trajectory that conform to a predetermined volume and/or trajectory. In general, the means for reducing uncontrolled electrostatic charge is selected according to the location, amount, and type of static electricity to be eliminated. Thus, for example, if a reservoir is prone to accumulate such uncontrolled electrostatic charge, the means for reducing uncontrolled electrostatic charge must be constructed according to the construction of the reservoir. Similarly, if a substrate onto which a droplet may be directed is susceptible to the accumulation of uncontrolled electrostatic charge, the means for reducing electrostatic charge may be constructed accordingly.
[0077] Typically, any effort to eliminate uncontrolled electrostatic charge may ensure that a droplet dispensed from the reservoir has a volume that does not deviate from the predetermined volume by more than about 10 %. Preferably, the droplet volume does not deviate from the predetermined volume by more than about 5 %. Optimally, the volume does not deviate from the predetermined volume by more than about 2 %. In addition, the trajectory of the droplet dispensed from the reservoir will typically not deviate from the predetermined trajectory by more than about 5°. Preferably, the trajectory does not deviate from the predetermined trajectory by more than about 1°. Optimally, the trajectory does not deviate from the predetermined trajectory by more than about 0.5°.
[0078] A number of electrostatic control techniques are known in the art and are suited for use with the present invention. Such techniques typically involve either addition or removal of electrons from the item that has accumulated uncontrolled electrostatic charge. On occasion, though, positive ions may be added or removed from the item. In general, electrostatic charge can be removed through grounding, induction, ionization, or a combination thereof. Such electrostatic charge neutralization may be effected immediately before or during the dispensation of a droplet.
[0079] Typically, uncontrolled electrostatic charge may be eliminated from an item through grounding, i.e., connecting the item via a conductor to an effectively infinite source of charge. Grounding is particularly suited for instances in which electrostatic charge is located in an ungrounded but highly conductive item. In such a case, the entire item may be neutralized when it is connected to ground at a single point. For example, items constructed from a material having a volume electrical resistivity of no more than about 104 ohm-cm and/or a surface electrical resistivity of no more than about 105 ohm/sq may be used. Preferably, the electrical resistivity is no more than about 103 ohm-cm and/or the surface electrical resistivity is no more than about 10 ohm/sq. For items comprised of a single material of high electrical resistivity, e.g., nonconductive polymers and ceramics, however, neutralization of the entire item may require the establishment of more than a single-point contact. In some instances, neutralization of an item may be achieved by providing the item with intermittent or sustained contact with an electrically conductive solid material.
[0080] Removing or neutralizing electrostatic charge by induction is a time-tested method suitable for use with any nonconductive material, insulated material, or ungrounded conductive material. Induction requires the use of an electrically conductive induction member that operates in a manner similar to the operation of a lightning rod. Typically, a grounded induction member, such as tinsel or a brush, is placed in close proximity, e.g., about 0.5 cm to about 1.0 cm, to the surface of the material to be neutralized. If the electrostatic charge on the material reaches or exceeds a threshold level, e.g., at least several thousand volts, the energy concentrated on the ends of the induction member will induce ionization. When the electrostatic charge is negative in polarity, positive ions from the grounded member will be attracted by the static laden surface. Conversely, if the static charge is positive in polarity, negative ions from the grounded member will be attracted back to the charged area.
[0081] It should be noted, however, that since a threshold voltage is required to "start" the process, induction may not reduce or neutralize static electricity to the ground potential level. In addition, an ungrounded induction member will remove charge for a short period of time only. Eventually the induction member will self charge and stop working when the electric field between the ends and the charged surface is reduced to a level that cannot support ionization. Thus, passive static control devices relying solely on induction tend to leave a residual charge.
[0082] Ionization techniques typically involve the production of both positive and negative ions to be attracted by the material to be neutralized. This may be achieved by generating an alternating electric field between a sharp point in close proximity to a grounded shield or casing. As the extremes of potential difference are reached, the air between the sharp point and the grounded casing is broken down. As a result, positive and negative ions are generated. In other words, half of the cycle is utilized to generate negative ions and the other half is utilized to generate positive ions. When a 60 Hz unit is employed, the polarity of ionization is changed every 1/120 of a second. If the material to be neutralized is positively charged, it will immediately absorb negative ions and repel the positive ions into space. Conversely, if the material to be neutralized is negatively charged, it will absorb the positive ions and repel the negative ions. When the material becomes neutralized, there is no longer electrostatic attraction and the material will cease to absorb ions.
[0083] Other equipment may also be used to generate ionized air for electrostatic neutralization. Nuclear-powered ionizers are known in the art. For example, Polonium 210 isotopes may be used to generate ions. Since Polonium has a half-life of only 138 days, such ionizers continually lose their strength and must be replaced annually. Similarly, electromagnetic radiation sources may be used to eliminate electrostatic charge. In some instances, such electromagnetic sources employ an ultraviolet radiation generator.
[0084] In some instances, surface conductivity of an item may be increased through the use of additives such anti-static sprays. An ordinary anti-static spray is comprised of a surfactant diluted in a solvent. A fire retardant may be added to counter the flammability of the solvent. Once applied to the surface of the item, the fire retardant and solvents evaporate, leaving a conductive coating on the surface of the material. The plastic has now become conductive and as long as this coating is not disturbed, it will be difficult to generate static electricity in this material. Thus, it should be evident that neutralization of an item may involve establishing intermittent or prolonged contacting of the item with a liquid and/or electrostatic-charge-reducing fluid. For example, when a fluid acoustic coupling medium is employed through which the ejector is acoustically coupled to the reservoir, the acoustic coupling medium may be comprised of an electrostatic-charge-reducing fluid.
[0085] Thus, it should be apparent that one of ordinary skill in the art may adapt any of the above-described or known equipment and techniques for reducing uncontrolled electrostatic charge for use with the present invention. It is also noted that use of a means for reducing uncontrolled electrostatic charge does not exclude the controlled use of ionization technology for directing droplet trajectory. Such technologies are generally well known in the art and are described, for example, in U.S. Patent Application Publication Nos. 20020109084 and 20020125424, each to Ellson et al. Because uncontrolled electrostatic charging may occur with the use of ionization technology to direct droplet trajectories, the invention may also be used to ensure that dispensed droplets conform to a predetermined size and/or predetermined trajectory.
[0086] However, it is generally preferred that all electric fields are eliminated with the practice of the invention. Thus, the invention preferably involves dispensing one or more droplets in the absence of any electrostatic charge or electric field that alters the trajectory and/or size of dispensed droplets. For example, in high-throughput and array applications, it is desirable to have control over the direction, volume, and velocity of dispensed droplets onto a droplet-receiving surface. Sometimes, production of a droplet of appropriate direction, volume, and velocity is accompanied by the production of a secondary or satellite droplet that should not be deposited onto the droplet-receiving surface. Using an electric field may accelerate both drops onto a receiving surface. In addition, electric fields may adversely interfere with droplet formation so as to result in difficulty in controlling droplet size.
[0087] FIG. 1 illustrates an exemplary focused acoustic ejection device suitable for use with the invention, in simplified cross-sectional view. As with all figures referenced herein, in which like parts are referenced by like numerals, FIG. 1 is not to scale, and certain dimensions may be exaggerated for clarity of presentation. The device 11 includes a plurality of reservoirs, i.e., at least two reservoirs— a first reservoir indicated at 13 and a second reservoir indicated at 15. Each reservoir may contain a combination of two or more immiscible fluids, and the individual fluids, and the fluid combinations in the different reservoirs may be the same or different. As shown, reservoir 13 contains fluid 14, and reservoir 15 contains fluid 16. Fluids 14 and 16 have fluid surfaces respectively indicated at 14S and 16S. As shown, the reservoirs are of substantially identical construction so as to be substantially acoustically indistinguishable, but identical construction is not a requirement. The reservoirs are shown as separate removable components but may, if desired, be fixed within a plate or other substrate. Each of the reservoirs 13 and 15 is axially symmetric as shown, having vertical walls 13W and 15W extending upward from circular reservoir bases 13B and 15B and terminating at openings 13O and 15O, respectively, although other reservoir shapes may be used. The material and thickness of each reservoir base should be such that acoustic radiation may be transmitted therethrough and into the fluid contained within the reservoirs. [0088] The device also includes an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation, and a focusing means 37 for focusing the acoustic radiation at a focal point near the fluid surface from which a droplet is to be ejected, wherein the focal point is selected so as to result in droplet ejection. When a plurality of immiscible fluid are used (not shown), the focal point may be in any layer. For example, when two immiscible fluids are used, the focal point may be in the upper fluid layer or in the lower fluid layer, e.g., just below the interface therebetween. As shown in FIG. 1, the focusing means 37 may comprise a single solid piece having a concave surface 39 for focusing acoustic radiation, but the focusing means may be constructed in other ways as discussed below. The acoustic ejector 33 is thus adapted to generate and focus acoustic radiation so as to eject a droplet of fluid from each of the fluid surfaces 14S and 16S when acoustically coupled to reservoirs 13 and 15, respectively. The acoustic radiation generator 35 and the focusing means 37 may function as a single unit controlled by a single controller, or they may be independently controlled, depending on the desired performance of the device. Typically, single ejector designs are preferred over multiple ejector designs, because accuracy of droplet placement, as well as consistency in droplet size and velocity, are more easily achieved with a single ejector.
[0089] Optimally, acoustic coupling is achieved between the ejector and each of the reservoirs through indirect contact. In FIG. 1 A, an acoustic coupling medium 25 is placed between the ejector 33 and the base 13B of reservoir 13, with the ejector and reservoir located at a predetermined distance from each other. The acoustic coupling medium 25 is introduced from a coupling medium source 27 via dispenser 29. Also as depicted in FIG. 1, an optional collector 47 is employed to collect coupling medium that may drip from the lower surface of either reservoir. As the collector 47 is depicted as containing the coupling medium source 27, it is evident that the coupling medium may be reused. Other means for introducing and/or placing the coupling medium, e.g., as described below, may be employed as well. By using an electrically conductive fluid as the acoustic coupling medium, the coupling medium source 27 and dispenser 29 serve as a means for reducing uncontrolled electrostatic charge from the reservoirs.
[0090] In operation, each reservoir 13 and 15 of the device is filled with different fluids, as explained above. The acoustic ejector 33 is positionable by means of ejector positioning means 61, shown below reservoir 13, in order to achieve acoustic coupling between the ejector and the reservoir through acoustic coupling medium 25. If droplet ejection onto a substrate is desired, a substrate 53 may be positioned above and in proximity to the first reservoir 13 such that one surface of the substrate, shown in FIG. 1 as underside surface 53S, faces the reservoir and is substantially parallel to the surface 14S of the fluid 14 therein. The substrate 53 is held by substrate positioning means 65, which, as shown, is grounded. Thus, when the substrate 53 is comprised of a conductive material, the substrate 53 is grounded as well. Once the ejector, the reservoir, and the substrate are in proper alignment, the acoustic radiation generator 35 is activated to produce acoustic radiation that is directed by the focusing means 37 to a focal point 14P near the fluid surface 14S of the first reservoir. As a result, droplet 14D is ejected from the fluid surface 14S, optionally onto a particular site (typically although not necessarily, a pre-selected, or "predetermined" site) on the underside surface 53S of the substrate. The ejected droplet may be retained on the substrate surface by solidifying thereon after contact; in such an embodiment, it is necessary to maintain the substrate surface at a low temperature, i.e., at a temperature that results in droplet solidification after contact. Alternatively, or in addition, a molecular moiety within the droplet attaches to the substrate surface after contact, through adsorption, physical immobilization, or covalent binding.
[0091] Then, as shown in FIG. IB, a substrate positioning means 65 may be used to reposition the substrate 53 (if used) over reservoir 15 in order to receive a droplet therefrom at a second site. FIG. IB also shows that the ejector 33 has been repositioned by the ejector positioning means 61 below reservoir 15 and in acoustically coupled relationship thereto by virtue of acoustic coupling medium 25. Once properly aligned, as shown in FIG. IB, the acoustic radiation generator 35 of ejector 33 is activated to produce acoustic radiation that is then directed by focusing means 37 to a focal point 16P within the reservoir fluids in reservoir 15, thereby ejecting droplet 15D, optionally onto the substrate.
[0092] It should be evident that such operation is illustrative of how the inventive device may be used to eject a plurality of droplets from reservoirs in order to form a pattern, e.g., an array, on the substrate surface 53S. It should be similarly evident that the device may be adapted to eject a plurality of droplets from one or more reservoirs onto the same site of the substrate surface. Furthermore, the ejection of a plurality of droplets may involve one or more ejectors. In some instances, the droplets are ejected successively from one or more reservoirs. In other instances, droplets are ejected simultaneously from different reservoirs.
[0093] As depicted in FIG. 2, the invention may be used with a single reservoir as well to improve the accuracy of droplet dispensation therefrom into an inlet opening of a sample vessel. Axially symmetric and grounded capillary 53 having an inlet opening 53O disposed on a terminus 53S thereof is provided as a sample vessel. Due to the axial symmetry of the capillary 53, the inlet opening 53O has a circular cross section. As such, the opening has a limiting dimension equal to its diameter.
[0094] A hemispherical volume of fluid 14 on a substantially flat surface 13S of a substrate 13 serves a reservoir. As shown, the substrate 13 is grounded so that it does not have any uncontrolled electrostatic charge. The shape of fluid 14 is a function of the sample wetting properties with respect to the substrate surface 13S. Thus, the shape can be modified with any of a number of surface modification techniques. In addition, an ejector 33 is provided comprising an acoustic radiation generator 35 for generating radiation, and a focusing means 37 for directing the radiation at a focal point 14P near the surface 14S of the fluid 14. The ejector 33 is shown in acoustic coupling relationship to the substrate 13 through coupling fluid 25. Proper control of acoustic wavelength and amplitude results in the ejection of a droplet 14D from the fluid 14 on the substrate 13. As the droplet 14D is shown having a diameter only slightly smaller than the diameter of the inlet opening 53O, it is evident that this configuration requires strict control over the droplet size and trajectory. Thus, the substrate is 13 grounded as well.
[0095] It should be noted that although the invention is well suited for use with any fluid, the influence of the uncontrolled electrostatic charge on droplet volume and/or trajectory is particularly pronounced with ionic compounds such as charged drug moieties. In addition, the presence of uncontrolled electric fields also tends to affect polar fluids with relatively high dielectric constants (k) such as water and dimethylsulfoxide (k = 80 and 48, respectively, at room temperature). As typical drug-screening compound libraries may contain compounds with varying polarities and dielectric constants, such libraries would be influenced differently by the same electrostatic charge. Thus, it should be evident that the invention is particularly suited for use in conjunction with fluidic manipulation associated with compound libraries. [0096] The invention may be suitable for use with any of the performance enhancing features associated with acoustic technologies such those described in U.S. Patent Application Publication No. 20030230344 to Ellson et al. relating to acoustic control of the composition and/or volume of fluid in a reservoir. In addition, the invention may be used in a number of contexts such as handling pathogenic fluids and manipulating cells and particles (see U.S. Patent Application Publication Nos. 20020090720 and 20020094582).
1
[0097] As discussed above, current positioning technology allows for an ejector positioning means to move from one reservoir to another quickly and in a controlled manner. For example, when a single acoustic radiation generator is used for ejecting and optionally assessing the contents of a plurality of reservoirs, the generator may be placed in acoustic coupling relationship in rapid succession to each of the reservoirs via the acoustic coupling fluid. Accordingly, the generator, the reservoirs, or both must be rapidly displaced with respect to each other for high-throughput techniques. Such rapid movement may cause uncontrolled flow of the acoustic coupling fluid. As a result, conformal contact between the acoustic generator and the reservoirs may not be achieved, thereby compromising the performance of the device. In some instances, uncontrolled acoustic fluid flow may result in the contamination of the reservoir contents, presence of sound-reflecting bubbles in the acoustic path, and/or degradation of device components.
[0098] In addition, when the an acoustic coupling medium is comprised of an electrostatic-charge-reducing fluid, nonconforrhal contact of the coupling medium to the reservoir may interfere with the dissipation of uncontrolled electrostatic charge on the reservoir. Thus, a means may be provided for eliminating uncontrolled flow of the acoustic coupling fluid at the exterior surface as a result of movement of the acoustic radiation generator. Such means may serve to facilitate and improve control over the elimination of electrostatic charge.
[0099] To elucidate the problems associated with uncontrolled flow of an acoustic coupling fluid, FIG. 3 depicts an acoustic device prone to such problems in simplified cross- sectional view. The device is shown in operation to form a biomolecular array bound to a substrate. The device 11 is generally similar to that depicted in FIG. 1 and includes two reservoirs, with a first reservoir indicated at 13 and a second reservoir indicated at 15. As shown, the first reservoir 13 contains a first fluid 14 and the second reservoir 15 contains a second fluid 16. Fluids 14 and 16 each have a fluid surface respectively indicated at 14S and 16S. As depicted, fluids 14 and 16 are of differing volumes and heights. That is, the distance between surface 14S and base 13B is greater than the distance between surface 16S and base 15B.
[0100] The device also includes an acoustic radiation generator 35 that contains a transducer 36, e.g., a piezoelectric element, commonly shared by an analyzer. As shown, a combination unit 38 is provided that both serves as a controller and a component of an analyzer. Operating as a controller, the combination unit 38 provides the piezoelectric element 36 with electrical energy that is converted into mechanical and acoustic energy. Operating as a component of an analyzer, the combination unit receives and analyzes electrical signals from the transducer. The electrical signals are produced as a result of the absorption and conversion of mechanical and acoustic energy by the transducer.
[0101] As shown in FIG. 3, the focusing means 37 is comprised of a single solid piece having a concave surface 39 for focusing acoustic radiation. The acoustic ejector 33 is thus adapted to generate and focus acoustic radiation so as to eject a droplet of fluid from each of the fluid surfaces 17 and 19 when acoustically coupled to reservoirs 13 and 15, and thus to fluids 14 and 16, respectively. The acoustic radiation generator 35 and the focusing means 37 function as a single unit controlled by a single controller.
[0102] In operation, as illustrated in FIG. 3A, a dispenser 29 places an acoustic coupling fluid 25 between the ejector 33 and the base 13B of reservoir 13, with the ejector placed at a predetermined distance from each the reservoir by positioning means 61. The dispenser 29 dispenses sufficient coupling fluid 25 so that the fluid established conformal contact between the concave surface 39 and base 13B. Once the ejector, the reservoir, and the substrate are in proper alignment, the acoustic radiation generator 35 is activated to produce acoustic radiation that is directed toward a free fluid surface 14S of the first reservoir. The acoustic radiation will then travel in a generally upward direction toward the free fluid surface 14S. The acoustic radiation will be reflected. By determining the time it takes for the acoustic radiation to be reflected by the fluid surface back to the acoustic radiation generator, and then correlating that time with the speed of sound in the fluid, the distance-and thus the fluid height-may be calculated. [0103] In order to form a biomolecular array on a substrate using the device, substrate
53 is positioned above and in proximity to the first reservoir 13 such that one surface of the substrate, shown in FIG. 3 as underside surface 51, faces the reservoir and is substantially parallel to the surface 14S of the fluid 14 therein. Due to the height of fluid 14, the ejector 33 is moved toward to the reservoir 13 to ensure that the focal point of the ejection acoustic wave is near the fluid surface 14S, where desired. That is, the ejector 33 is moved positively along axis Z. As a result, acoustic coupling fluid 25 is displaced through uncontrollable flow. When movement of the ejector is at a high velocity, the acoustic coupling fluid may be squirted or sprayed in a direction perpendicular to axis Z.
[0104] In any case, once the ejector, the reservoir, and the substrate are in proper alignment, the acoustic radiation generator 35 is activated to produce acoustic radiation that is directed by the focusing means 37 to a focal point 14P near the fluid surface 14S of the first reservoir. That is, an ejection acoustic wave having a focal point near the fluid surface is generated in order to eject at least one droplet of the fluid. As a result, droplet 14D is ejected from the fluid surface 14S onto a designated site on the underside surface 51 of the substrate.
[0105] Then, as shown in FIG. 3C, a substrate positioning means 65 repositions the substrate 53 over reservoir 15 in order to receive a droplet therefrom at a second designated site. FIG. IC also shows that the ejector 33 has been repositioned by the ejector positioning means 61 below reservoir 15 and in acoustically coupled relationship thereto by virtue of acoustic coupling fluid 25. Again, the dispenser 29 dispenses sufficient coupling fluid 25 so that the fluid establishes conformal contact between the concave surface 39 and base 15B. Once properly aligned, the acoustic radiation generator 35 of ejector 33 is activated to produce low energy acoustic radiation to assess the height of fluid 16 in reservoir 15 and to determine whether and/or how to eject fluid from the reservoir.
[0106] Due to the height of fluid 16, the ejector 33 is moved away from the reservoir
15 to ensure that the focal point of the ejection acoustic wave is near the fluid surface 16S, where desired. That is, the ejector 33 is moved negatively along axis Z. As a result, acoustic coupling fluid 25 flows uncontrollably so that it no longer conforms to surface 39 and base 15B. In some instances, bubbles will form within the acoustic coupling fluid. For example, air bubbles may be sucked into fluid. Under extreme circumstances, bubbles may be formed as a result of cavitation. Thus, any droplet 16D ejected from reservoir 15 toward substrate 53 may be misdirected due to the lack of conformal contact.
[0107] Thus, it should be apparent that uncontrolled flow of the coupling fluid is particularly problematic when the acoustic generator in contact with the coupling fluid is moved rapidly relative to the exterior surface. Correspondingly, when acoustic ejection and/or assessment techniques are carried out that involves use of a single acoustic generator rapidly and successively coupled via an acoustic coupling fluid to a plurality of reservoirs, uncontrolled fluid flow may compromise the viability of the techniques, particularly in the context of high-throughput combinatorial methods.
[0108] Accordingly, another embodiment of the invention provides an acoustic device as described above that also includes a means for delivering an acoustic coupling fluid to an exterior surface of the reservoir and a means for eliminating uncontrolled flow of the acoustic coupling fluid at the exterior surface as a result of movement of the acoustic radiation generator.
[0109] Although any of a number of different means may be used to deliver the acoustic coupling fluid to the exterior surface of the reservoir, such means typically includes a source of the acoustic coupling fluid in fluid communication with a nozzle having an outlet that opens toward the exterior surface of the reservoir. Often, the acoustic coupling fluid is comprised of water. However, fluids similar to water may be used as well. For example, if the device is constructed for operation with water as an acoustic coupling fluid, the acoustic coupling medium may be comprised of a fluid that exhibits an attenuation coefficient for acoustic radiation of a selected frequency similar to that of water. The selected frequency is typically the operating frequency of the device. For example, if a particular frequency is found to be the optimal frequency for droplet ejection, that frequency may be the selected frequency associated with the attenuation coefficient. Typically, the coupling fluid exhibits an attenuation coefficient for acoustic radiation of a selected frequency that differs from the attenuation coefficient of water at the same frequency by no more than about 10%. Preferably, the difference in attenuation coefficient is no more than about 5%. Optimally, the difference in attenuation coefficient is no more than about 1%. In any case, one of ordinary skill in the art will recognize that fluids that exhibit a lower degree of acoustic attenuation than water may be advantageously used to reduce the power for acoustic radiation generation. In addition, the acoustic coupling fluid is typically directed to flow from the source to the outlet at a rate sufficient for the acoustic coupling fluid to establish conformal contact with the exterior surface of the reservoir.
[0110] The device may include a collector as well as a means for positioning the nozzle. The collector is placed in fluid-receiving relationship to the exterior surface of the reservoir so as to collect excess acoustic coupling fluid flowing therefrom. For example, the nozzle may be placed directly below the exterior surface of the reservoir such that acoustic coupling fluid emerging from the nozzle is directed upward for conformal contact with the exterior surface of the reservoir. To allow facile collection of the acoustic coupling fluid flowing downward from the exterior reservoir surface, the nozzle may be located within the collector.
[0111] The nozzle is typically placed no closer than a predetermined distance from the exterior surface of the reservoir so as to avoid contact between the nozzle and the surface. In addition, some embodiments allow acoustic radiation is propagated through the acoustic coupling fluid in the nozzle and the exterior surface into the reservoir. Thus, a particularly useful design allows the nozzle and the acoustic radiation generator to move along the same axis extending from the exterior surface of the reservoir. Typically, the axis is perpendicular to the exterior surface.
[0112] FIG. 4 depicts an exemplary acoustic device having a nozzle and collector as described above. As shown, a single reservoir 13 containing a fluid 14 having a fluid surface indicated at 14S. Reservoir 13 has a base indicated at 13B and an opening indicated at 13O. Dispenser 29 provided is comprised of a nozzle 30 that terminates upwardly at an outlet 32 directed toward the reservoir base 13B and downwardly at a pump 34 for pumping acoustic coupling fluid 25 upwardly through the nozzle 30. Located within the nozzle 30 is an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation and a focusing means 37 for focusing the acoustic radiation at a focal point within the fluid 14P from which a droplet is to be ejected, near the fluid surface 14S. Positioning means 61 serves to controllably move ejector 33 within nozzle 30 along axis Z. The device also includes a collector 31 for collecting coupling fluid that flows from base 13B. As shown, the nozzle 30 is located within the collector 31. Located at the bottom of the collector 31 and in fluid communication with the pump 34 is a source 27 of acoustic coupling fluid.
[0113] In operation, positioning means 70 positions the dispenser 29 at predetermined distance to the reservoir base 13B. The pump 34 draws acoustic coupling fluid from the source 27 and forces the acoustic coupling fluid upward through the nozzle 30. The flow of acoustic coupling emerging from outlet 32 is typically maintained at constant rate and sufficiently high to allow the coupling fluid to establish and maintain conformal contact with reservoir base 13B. After contact with reservoir base 13B, acoustic coupling fluid falls back down into collector 31, where the coupling fluid redirected toward source 27 and pump 34 for reuse.
[0114] At a constant flow rate, the acoustic coupling fluid 25 between the ejector 33 and the base 13B allows for acoustic radiation generated by the generator 35 to be transmitted therethrough. As a result, acoustic radiation will then travel in a generally upward direction, through base 13B and fluid 14 toward the free fluid surface 14S. The acoustic radiation reflected by free surface 14S may then be analyzed. If needed to ensure that the acoustic radiation is focused near the fluid surface 14S to eject a droplet therefrom, positioning means in the form of telescoping rod 61 may be employed to move ejector 33 to an appropriate location within nozzle 30. For example, the rod 61 may be adapted to elongate in a telescoping manner within the nozzle to move ejector 33 toward the outlet 32. Similarly, the ejector 33 is moved toward pump 34 when rod 61 is retracted. In any case, the ejector 33 may be maintained at a fixed distance from the fluid surface 14S so as to ensure that the acoustic radiation remains focused near the fluid surface 14S as the fluid level in the reservoir 13 is lowered due to the ejection of droplets therefrom.
[0115] A number of different designs and mechanisms may be used as a means for eliminating uncontrolled flow of the acoustic coupling fluid. For example, when a nozzle as depicted in FIG. 4 is employed, uncontrolled fluid flow may be avoided simply by immobilizing the relative positions of the reservoir 13 and the nozzle 30 and maintaining fluid flow from outlet 32 at a constant rate. Nevertheless, it should be apparent that any movement of ejector 33 within nozzle 30, particularly rapid movement, may disturb the rate of fluid flow from outlet 32, particularly when the pump 34 moves acoustic fluid at a constant rate. For example, as ejector 33 is moved upward toward outlet 32, the rate of fluid flow emerging from outlet 32 will tend to increase temporarily as rod 61 displaces as coupling fluid within the nozzle 30. Similarly, movement of ejector 33 downward toward pump 34 will cause the rate of fluid flow emerging from outlet 32 to decrease.
[0116] Thus, means for eliminating uncontrolled coupling fluid flow from outlet 32, may serve to maintain the fluid pressure at outlet 32 at a constant level. For example, the means for positioning the nozzle and the means for positioning the generator may be synchronized to maintain flow of acoustic coupling fluid from the nozzle at a constant rate, thereby serving as the means for eliminating uncontrolled flow. In addition, a displacement member that maintains the acoustic coupling fluid at a constant volume within the nozzle may be used in response any movement of the acoustic radiation generator within the nozzle. Such displacement members may be selected from pistons, diaphragm, combinations thereof, and other mechanisms. In some instances, the displacement member may be at least partially located within the nozzle. In addition or in the alternative, the displacement member may be at least partially located external to the nozzle in a chamber that fluidly communicates with the nozzle.
[0117] A flow rate regulator may be advantageously used to adjust the flow rate of the acoustic coupling fluid from the source to the outlet according to movement of the acoustic radiation generator within the nozzle. For example, an adjustable valve may be provided downstream from the source and upstream from the outlet to adjust the flow rate of the acoustic coupling fluid. Flow rate regulator technology is well known in the art and one of ordinary skill should be able to adapt the device to incorporate such regulators.
[0118] As discussed above, the means for positioning the acoustic radiation generator may sometime cause uncontrolled flow of the acoustic fluid. Thus, in some instances, an acoustic radiation generator positioning means may be used that has a structure does not substantially alter the volume of the acoustic coupling fluid within the container while positioning the acoustic radiation generator. In such instances, the structure itself serves as the means for eliminating uncontrolled flow of the acoustic coupling fluid. FIG. 5 depicts an exemplary acoustic device that employs a stationary opposing piston design to maintain coupling fluid flow at a constant rate from a nozzle outlet. The opposing piston design operates by maintaining the acoustic coupling fluid at a constant volume within the nozzle. Dispenser 29 provided is comprised of a nozzle 30 that terminates upwardly at an outlet 32 directed upwardly for delivering acoustic coupling fluid 25 to the exterior surface of a reservoir (not shown). Located within the nozzle 30 is an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation and a focusing means 37 for focusing the acoustic radiation. Positioning means in the form of a platform 61 serves to move ejector 33 within nozzle 30 along axis Z in a controlled manner. Also provided is a stationary piston 72 that extends through the nozzle 30 and into a corresponding opening 74 in platform 61. As depicted, the volume of acoustic coupling fluid 25 within the nozzle remains constant as ejector 33 is moved along axis Z as long as piston 72 extends through opening 74.
[0119] In some instances, a means other than a nozzle may be used to deliver acoustic coupling fluid to the exterior surface of a reservoir. For example, a container may be sealed against the reservoir and filled with the acoustic coupling fluid such that the acoustic coupling fluid is in conformal contact with the exterior surface of the reservoir. In such a case, the acoustic radiation generator may be movable within the container.
[0120] FIG. 6 depicts an acoustic device similar to that depicted in FIG. 3 with some notable differences relating to the means for eliminating uncontrolled coupling fluid flow. The device 11 includes two attached reservoirs provided in the form of wells 13 and 15 of a well plate 12. The wells 13 and 15 share a common underside surface 12B that is substantially planar. Like the device of FIG. 3, the device of FIG. 6 also includes an acoustic ejector 33 comprised of an acoustic radiation generator 35 for generating acoustic radiation and a focusing means 37 for focusing the acoustic radiation at a focal point within the fluid from which a droplet is to be ejected, near the fluid surface. A positioning means 61 serves to couple the ejector 33 successively to each of the wells.
[0121] Unlike the device depicted in FIG. 3, the dispenser is replaced with a container
29 having a base and walls extending upward from the base and terminating at an opening 29O. Completely filled with coupling fluid 25, the container 29 is positioned such that the opening 29O contacts with the underside surface 12B of the well plate 12, thereby forming a seal therebetween. As a result, acoustic ejector 33 and positioning means 61 are both sealed within the container 29 and submerged in coupling fluid 25. Because the volume of the coupling fluid remains 25 unaltered irrespective of the movement and or positioning of the ejector 33 within the container 29, uncontrolled flow of the acoustic coupling fluid at the exterior surface as a result of movement of the acoustic radiation generator is eliminated.
[0122] Alternatively, when the movement of the acoustic generator 35 is accompanied by displacement of volume in the container 29, any of the above-described means for eliminating uncontrolled coupling fluid flow associated with the nozzle may be used with the container as well.
[0123] From the above, it should be apparent that a method is provided for transmitting acoustic radiation into a reservoir. The method involves simultaneously delivering an acoustic coupling fluid to an exterior surface of a reservoir adapted to contain a fluid and positioning an acoustic radiation generator for generating acoustic radiation in acoustic coupling relationship via the acoustic coupling fluid to the reservoir. This is carried out in a manner that avoids uncontrolled flow of the acoustic coupling fluid at the exterior surface. Once the acoustic radiation generator is in position, it is activated so as to generate and transmit acoustic radiation through the exterior surface and into any fluid contained in the reservoir.
[0124] Variations of the present invention will be apparent to those of ordinary skill in the art. For example, any of a number of positioning means known in the art may used with the invention. Such positioning means may be constructed from, e.g., levers, pulleys, gears, a combination thereof, or other mechanical means known to one of ordinary skill in the art. In addition, as alluded to above, positioning means may be used to move items such as the reservoir, the acoustic generator, the coupling fluid delivering means, or a combination thereof, to provide relative motion therebetween. One of ordinary skill in the art will recognize that relative motion may be provided by holding any one or a combination of the items in a fixed position while the allowing the positioning means to move the remaining items. Furthermore, while the invention has been described above in the context of single- element acoustic generator, multiple element acoustic radiation generators such as transducer assemblies may be used as well. That is, linear acoustic arrays, curvilinear acoustic arrays, annular acoustic arrays, phased acoustic arrays, and other transducer assemblies may be used in conjunction with the invention as well. Moreover, since acoustic detectors, like acoustic generators, may be used in conjunction with acoustic coupling fluids, those of ordinary skill in the art will be able to substitute acoustic detectors in place of acoustic generators in certain applications.
[0125] It is to be understood that while the invention has been described in conjunction with the preferred specific embodiments thereof, that the foregoing description and the examples that follow are intended to illustrate and not limit the scope of the invention. Other aspects, advantages, and modifications within the scope of the invention will be apparent to those skilled in the art to which the invention pertains.
[0126] The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how to implement the invention, and are not intended to limit the scope of what the inventors regard as their invention.
EXAMPLE 1
A solution containing 70% by volume dimethylsulfoxide and 30% by volume water was placed within each well of a polystyrene well plate containing 384 substantially identical wells. An acoustic ejector having an F2 lens that served to focus acoustic radiation was placed in acoustic coupling relationship successively with each reservoir in substantially the same manner. Without removing uncontrolled electrostatic charge from the well plate, acoustic radiation having a frequency of 10 MHz was directed by the F2 lens into each reservoir so as to eject at least one droplet from each well. In some instances, secondary or satellite droplets were produced in addition to the primary droplets. The primary droplets exhibited a volume variation of over 25% as well as variations in trajectory.
EXAMPLE 2
Each well of the same polystyrene well plate described in Example 1 was again filled with a solution containing 70% by volume dimethylsulfoxide and 30% by volume water. However, uncontrolled electrostatic charge was removed from the well plate using an ionizer before the acoustic ejector was placed in acoustic coupling relationship successively with each reservoir. Acoustic radiation of having a frequency of 10 MHz was again directed by the F2 lens into each reservoir so as to eject at least one droplet from each well. No secondary or satellite droplets were produced. The primary droplets exhibited a volume variation of less than about 2%. No variations in the trajectory of the droplets were observed.

Claims

CLAIMS:
1. In a device comprised of a reservoir adapted to contain a fluid and a dispenser for dispensing a fluid droplet from the reservoir, the improvement comprising employing a means for reducing uncontrolled electrostatic charge on the reservoir when the reservoir is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom, wherein the means for reducing uncontrolled electrostatic charge is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
2. In a device comprised of a reservoir adapted to contain a fluid, a dispenser for dispensing a fluid droplet from the reservoir, and a substrate positioned to receive the dispensed droplet, the improvement comprises employing a means for reducing uncontrolled electrostatic charge on the substrate when the substrate is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of the dispensed droplet, wherein the means for reducing uncontrolled electrostatic charge is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
3. A device for acoustically ejecting a droplet of fluid from a reservoir, comprising: a reservoir adapted to contain a fluid; an ejector for ejecting a droplet from the reservoir, comprising an acoustic radiation generator for generating acoustic radiation and a focusing means for focusing the acoustic radiation generated; and a means for positioning the ejector in acoustic coupling relationship to the reservoir; and a means for reducing any uncontrolled electrostatic charge on the device or a portion thereof that alters the volume and/or trajectory of a droplet ejected from the reservoir, wherein the means for reducing uncontrolled electrostatic charge is effective to ensure that the volume and/or trajectory of the ejected droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
4. The device of any one of claims 1-3, wherein accumulation of electrostatic charge uncontrollably alters the volume of a droplet dispensed from the reservoir.
5. The device of claim 4, wherein the means for reducing uncontrolled electrostatic charge is effective to ensure that a droplet dispensed from the reservoir has a volume that does not deviate from the predetermined volume by more than about 10 %.
6. The device of any one of claims 1-3, wherein the reservoir is prone to accumulate electrostatic charge that uncontrollably alters the trajectory of a droplet dispensed therefrom.
7. The device of claim 6, wherein the means for reducing uncontrolled electrostatic charge is effective to ensure that a droplet dispensed from the reservoir has a trajectory that does not deviate from the predetermined trajectory by more than about 5°.
8. The device of claim 1 or 2, wherein the dispenser is comprised of an ejector that does not require contact with a fluid in a reservoir to eject the fluid from the reservoir.
9. The device of claim 8, wherein the ejector is an acoustic ejector.
10. The device of claim 9, further comprising an acoustic coupling medium through which the ejector is acoustically coupled to the reservoir
11. The device of claim 3 or 10, comprising a single ejector.
12. The device of claim 11, wherein the reservoir is detachable from the device.
13. The device of claim 11, wherein the reservoir is comprised of a material having a volume electrical resistivity of at least 1013 ohm-cm and/or has a surface electrical resistivity of at least 1014 ohm/sq.
14. The device of claim 11, wherein the reservoir is comprised of a polymeric material.
15. The device of claim 11, comprising a plurality of reservoirs, each adapted to contain a fluid, wherein the means for reducing uncontrolled electrostatic charge reduces uncontrolled electrostatic charge on each of the reservoirs.
16. The device of claim 15, wherein the reservoirs are arranged in an array.
17. The device of claim 16, wherein the reservoirs are arranged in a rectilinear array.
18. The device of claim 15, wherein each reservoir is a well in a well plate.
19. The device of claim 11, wherein the focusing means exhibits an F-number of at least about 1.
20. The device of claim 19, wherein the focusing means exhibits an F-number of at least about 2.
21. The device of claim 20, further comprising a means for positioning the ejector in successive acoustic coupling relationship to each reservoir.
22. The device of claim 10, wherein the means for reducing uncontrolled electrostatic charge is comprised of the acoustic coupling medium, and the acoustic coupling medium is comprised of an electrostatic-charge-reducing fluid.
23. The device of claim 11 , wherein the means for reducing uncontrolled electrostatic charge comprises an electromagnetic radiation source, an electrically conductive solid material, an electrostatic-charge-reducing fluid, and/or an electrostatic-charge-reducing gas.
24. The device of claim 11, wherein the means for reducing uncontrolled electrostatic charge removes electrons, adds electrons, grounds uncontrolled electrostatic charge, operates through induction, and/or operates through ionization.
25. A device for acoustically ejecting a droplet of fluid from a reservoir, comprising: a reservoir adapted to contain a fluid; an ejector for ejecting a droplet from the reservoir, comprising an acoustic radiation generator for generating acoustic radiation and a focusing means for focusing the acoustic radiation generated; a means for positioning the ejector in acoustic coupling relationship to the reservoir; and an optional substrate positioned to receive the ejected droplet, wherein the reservoir, the optional substrate, or both are grounded and comprised of a material that either has an electrical resistivity of no more than about 1011 ohm-cm, has a
19 surface electrical resistivity of no more than about 10 ohm/sq, or both.
26. A device for containing one or more fluids, comprising: a well plate having an array of at least 96 substantially identical wells, each well adapted to contain a fluid, wherein the well plate is comprised of a material that is at least partially nonmetallic and either has an electrical resistivity of no more than about 1011 ohm- cm, has a surface electrical resistivity of no more than about 1012 ohm/sq, or both.
27. The device of claim 25 or 26, wherein the material is coated with an electrically conductive film.
28. The device of claim 27, wherein the electrically conductive film is metallic.
29. The device of claim 25 or 26, wherein the material contains an electrically conductive filler.
30. The device of claim 29, wherein the electrically conductive filler is carbon.
31. The device of claim 25 or 26, wherein the material is at least partially polymeric.
32. The device of claim 31, wherein the material is entirely polymeric.
33. The device of claim 31, wherein the polymeric material is selected from the group consisting of polyethylenes, polypropylenes, polybutylenes, polystyrenes, cyclic olefins, polythiophenes, polyacetylene, derivatives thereof, combinations thereof, and copolymers of any of the foregoing.
34. The device of claim 25 or 26, wherein the material is comprised of an electrically conductive polymer.
35. In a method for dispensing a droplet from a reservoir containing a fluid, the improvement comprises reducing uncontrolled electrostatic charge on the reservoir when the reservoir is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom, wherein uncontrolled electrostatic charge is reduced to a level effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
36. In a method for dispensing a droplet from a reservoir containing a fluid on to a substrate, the improvement comprises reducing uncontrolled electrostatic charge on the reservoir and/or the substrate when the reservoir and/or substrate is prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of the dispensed droplet, wherein the reduction of uncontrolled electrostatic charge is effective to ensure that the volume and/or trajectory of the dispensed droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
37. A method for acoustically ejecting a droplet of fluid from a reservoir, comprising: applying focused acoustic radiation in a manner effective to eject a droplet of fluid from the reservoir; and reducing any uncontrolled electrostatic charge that alters the volume and/or trajectory of the droplet ejected from the reservoir so as to ensure that the volume and/or trajectory of the ejected droplet do not substantially deviate from a predetermined volume and/or predetermined trajectory.
38. The method of any one of claims 35-37, wherein the uncontrolled electrostatic charge is reduced to a level effective to ensure that a droplet dispensed from the reservoir has a volume that does not deviate from the predetermined volume by more than about 10 %.
39. The method of any one of claims 35-37, wherein the uncontrolled electrostatic charge is reduced to a level effective to ensure that a droplet dispensed from the reservoir has a trajectory that does not deviate from the predetermined trajectory by more than about 5°.
40. The method of claim 35 or 36, wherein focused acoustic radiation is applied in a manner effective to eject a droplet of fluid from the reservoir.
41. The method of claim 37 or 40, wherein each of a plurality of droplets is successively ejected from the reservoir.
42. The method of any of claims 35-37, wherein a droplet is ejected from each of a plurality of reservoirs by applying focused acoustic radiation in a manner effective to eject a droplet of fluid from each of the reservoirs, wherein the uncontrolled electrostatic charge is reduced for each reservoir prone to accumulate uncontrolled electrostatic charge that alters the volume and/or trajectory of a droplet dispensed therefrom.
43. The method of any one of claims 37 and 40-42, wherein the uncontrolled electrostatic charge is reduced immediately before droplet ejection.
44. The method of any one of claim 37 and 40-42, wherein the uncontrolled electrostatic charge is reduced during droplet ejection.
45. The method of claim 37 or 40, wherein the focused acoustic radiation is applied through an acoustic coupling medium in contact with the reservoir and comprised of a electrostatic-charge reducing fluid.
46. The method of any one of claims 35-37, wherein the uncontrolled electrostatic charge is reduced by via application of radiation, contact with an electrically conductive solid material, contact with an electrostatic-charge-reducing fluid, and/or contacting with an electrostatic-charge-reducing gas.
47. The method of any one of claims 35-37, wherein the uncontrolled electrostatic charge is reduced by removing electrons, adding electrons, grounding the uncontrolled electrostatic charge, effecting electrostatic induction, and/or effecting ionization.
PCT/US2004/000522 2003-01-09 2004-01-09 Droplet dispensation from a reservoir with reduction in uncontrolled electrostatic charge WO2004063029A2 (en)

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AT04701230T ATE554931T1 (en) 2003-01-09 2004-01-09 DROPLETS DISPENSING DEVICE FROM A CONTAINER WITH UNCONTROLLED ELECTROSTATIC CHARGE REDUCTION
EP04701230A EP1585636B1 (en) 2003-01-09 2004-01-09 Droplet dispensation from a reservoir with reduction in uncontrolled electrostatic charge

Applications Claiming Priority (4)

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US10/340,557 US7070260B2 (en) 2003-01-09 2003-01-09 Droplet dispensation from a reservoir with reduction in uncontrolled electrostatic charge
US10/340,557 2003-01-09
US10/668,534 US6916083B2 (en) 2003-09-22 2003-09-22 Control over flow of an acoustic coupling fluid
US10/668,534 2003-09-22

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WO2004063029A3 WO2004063029A3 (en) 2004-11-18

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ATE554931T1 (en) 2012-05-15
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WO2004063029A3 (en) 2004-11-18

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