US 3813031 A
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United States Patent [191 Anderson 11] 3,813,031 [451 May 28,1974
[ ROTOR HAVING SAMPLE HOLDING 22 Filed: Aug. 2, 1972 211 App]. No.: 277,192
 US. Cl. 233/26  Int. Cl B04b 9/12  Field of Search. 233/26, 1 R, 27, 28; 356/197,
 References Cited .UNITED STATES PATENTS 3,326,458 6/1967 Meryman et a1 233/26 X 3,547,547 12/1970 Anderson 3,586,484 6/l971 Anderson 233/1 R Primary ExaminerGeorge H. Krizmanich Attorney, Agent, or FirmRoland A. Anderson  ABSTRACT A fast photometric analyzer of the multistation, rotary type is provided which is suitable for performing sample and reagent division, mixing, sedimentation, washing, transfer, and light absorbancy functions. Two sets of cavities are disposed in concentric annular arrays within a disk adapted to rotate about its central axis. The radially outer set of cavities constitutes a rotary system of cuvettes suitable for photometric measurements. Cavities in the radially inner set of cavities, herein designated loading cavities, communicate, on a one-to-one basis, with respective cuvette cavities. A sloping barrier is provided between each loading cavity and corresponding cuvette to allow for centrifugal transfer of fluids from the loading cavity to the cuvette upon rotation of the disk while preventing such transfer or return of fluids under static conditions. An opening is provided through each barrier to enhance mixing in the cuvettes. A vacuum activated drainline extends radially inward from a point adjacent the radial extremity of each cuvette to a vacuum source for removing overlaying liquid from the cuvettes. A central receiving and distribution chamber bordered by segmentation serrations is in fluid communication with each loading cavity to facilitate dynamic or static loading of those cavities.
4 Claims, 8 Drawing Figures EATENTEDHAY 28 1974 13.8 111031 sum 1 or 2 PHOTO DETECTOR r /1 BACKGROUND OF THE INVENTION The invention relates generally to photometric analyzers of the multistation, rotary type and more particularly to an improved photometric analyzer which is amenable to serological testing requiring repeated washing or dilution of particulates. It was made in the course of, or under, a contract with the U. S. Atomic Energy Commission.
Many serological tests involve phenomena shown by liquid-suspended particles having specific antigens or antibodies affixed thereto. In basic examples, these tests involve blood cells, bacterial cells, and the like. In more recently developed tests such as those described in copending application Ser. No. 223,018 of common assignee, the serological factors are attached to chemically stable carrier particles such as synthetic resin beads. Attachment of additional antibodies or antigens to these carrier particles may result in an agglutination reaction which may be observed directly. In other cases, reacted carrier particles may be centrifugally separated from the suspending liquid by sedimentation with or without a liquid density gradient, and the serological factor secondarily attached thereto identified by a change in particle sedimentation rate or banding density.
Most serological tests of the foregoing type require centrifugal sedimenation of the carrier particles and repeated washing steps. The actual means by which the various reactions are performed, the specific reagents used, the reaction times, and the sequence of the wahsing and reaction steps vary considerably for different tests. The unique parallel mode of operation of fast photometric analyzers of the rotary cuvette type, wherein samples and reagents are mixed in a centrifugal field and absorbance or fluorescent measurements are made on a multiplicity of reaction mixtures during rotation, makes them of interest for use in the automation of immunological analyses. This necessitates the development of a cuvette rotor which permits extensive washing of red cells or other carrier particles, reaction with one or more reagents (with interspersed washes, if necessary), and determination of the degree of agglutination, extent of hemolysis, and presence or absence of fluorescence-labelled antibody or radio-activity in either the supernatant or sediment.
It is, accordingly, a general object of the invention to provide a fast photometric analyzer of the rotary cuvette typw which is suitable for immunological analyses requiring repeated washings of carrier particles.
Another, more particular object of the invention is to provide a fast photometric analyzer of the rotary cuvette type which is capable of performing sample and reagent division, mixing, sedimentation, washing, transfer, and light absorbancy functions.
Other objects of the invention will be apparent from an examination of the following description of a preferred embodiment and the appended claims.
SUMMARY OF THE INVENTION In accordance with the invention, a photometric ana- I lyzer of the multistation, rotary cuvette type is provided which is capable of performing sample and reagent dirotate about its central axis. The radially outer set of cavities constitutes a rotary cuvette system where photometric measurements are made. Cavities in the radially inner set of cavities, herein designated loading cavities, communicate, on a one-to-one basis, with respective cavities in the radially outer set. A barrier between each loading cavity and a corresponding cuvette'allows centrifugal transfer of fluids upon rotation of the disk while preventing such transfer or return of fluids under static conditions. A radially extending passageway positioned angularly to one side of the barrier results in improved mixing in the cuvettes. A drainline extends radially inward from a point adjacent the radial extremity of each cuvette to a vacuum source for removing overlaying liquid within the cuvettes following particle sedimentation therein. A central receiving and distribution chamber bordered by segmentation serrations is in fluid communication with each loading cavity to facilitate loading of those cavities under static or dynamic conditions. Use of the subject analyzer permits the making of multiple serological analyses requiring repeated washing or dilution and sedimentation steps in a rapid manner.
BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a vertical sectional view, highly schematic, of a multistation, rotary type photometric analyzer made in accordance with the invention.
FIG. 2 is a top plan view of a rotor assembly used in the photometric analyzer of FIG. 1.
FIGS. 3, 4, 5, 6, 7, and 8 illustrate the operation of the rotor assembly of FIG. 2 in performing reaction, sedimentation, washing, and fluid transfer functions.
DESCRIPTION OF THE PREFERRED EMBODIMENT FIG. 1 illustrates, in a vertical view, a photometric analyzer made in accordance with the invention. A pancake-shaped rotor assembly 1 includes a flanged steel rotor body 2, transparent glass or plastic plates 3 and 4, a polytetrafluorethylene or other plastic disk 5 defining loading cavities and cuvettes, and a bolted flange ring 6 for holding plates 3 and 4 and disk 5 within rotor body 2. Plates 3 and 4 and disk 5 are sandwiched together between rotor body 2 and flange ring 6 to form loading cavities 7 and cuvettes 8 which are disposed in concentric annular arrays at respectively increasing rotor radii. Drainlines 9 extend from a point adjacent the radial extremity of each cuvette 8 to a central vacuum plenum 10 which passes through drive and support shaft 11 to a vacuum source (not shown). A sloping barrier 12 separates each loading cavity 7 from a corresponding cuvette 8 under static conditions while permitting centrifugal transfer of liquids from the loading cavities to the cuvettes. Recesses 13 in plate 4 are positioned directly above each barrier 12 to permit liquid to flow over the barrier into cuvettes 8 during rotation of the rotor assembly. The slope, spacing, and orientation of loading cavities 7, cuvettes 8, and drainlines 9 are further illustrated in the plan view of FIG. 2, which is described below.
Cuvettes 8 are designed to permit simultaneous photometric analyses of a multiplicity of discrete samples. To facilitate such analyses, a multiplicity of spacedapart holes 14, axially aligned with and equal in number to cuvettes 8, are provided in rotor body 2. Light passing through holes 14 continues through transparent plate 3, cuvettes 8, and transparent plate 4 to a photodetector as indicated schematically by a broken line in FIG. 1. A system of photometric light source, photodetecting means, and other ancillary electronic components suitable for use with the above rotor assembly is described in copending application of common applicant and assignee Ser. No. 784,739. That system will not be described here inasmuch as the invention in this case is an improvement restricted to the design of the ,rotor assembly and its method of use only and does not encompass the associated components described in the copending application.
Referring now to FIG. 2 where a plan view of disk 5 is shown, only two of sixteen pairs of loading cavities and cuvettes are shown for purposes of drawing simplification. A central liquid distribution chamber is provided with a serrated periphery in the form of stream segmenting blades 16 for dividing liquid inserted during rotation into substantially equal volumes and for distributing the volumes thus divided to respective loading cavities and cuvettes. As shown, radially extending passageways 17 are positioned angularly to one side of each barrier 12 to permit passage of liquid along the corresponding side of mixing cavities 7 and cuvettes 8, thereby enhancing the mixing action in cuvettes 8 through a vortex flow effect illustrated in FIG. 5. The operation of loading cavities 7, cuvettes 8, barrier 12, passageway 17, and drainlines 9 may best be understood by referring to FIGS. 3 through 8 where the stepwise operation of the subject photometric analyzer is illustrated.
An initial loading of blood samples or other particulate suspension 18 is first made under static conditions as illustrated in FIG. 3. Access to loading cavities 8 may be made through opening 19 in plate 4 (see FIG. 1) by any suitable loading apparatus such as a hypodermic syringe. Alternatively, a small transfer disk (not shown) may be used to perform the sample loading operation. FIG. 4 illustrates the centrifugal transfer of sample 18 to the radial extremity of a corresponding cuvette 8. Transfer ofv the sample to cuvette 8 is accomplished by the sample passing over the sloping barrier 12 through the clearances provided by recesses 13 in plate 4. As shown, the samples are spread in a thin layer on the radially outer wall 20 of each cuvette 8 by the centrifugal action.
FIG. 5 shows a diluent stream being directed against segmentation blades 16. After striking plates 16, the divided diluent flows along one side of each loading cavity 7, through passageway 17, and thence into cuvette 8 where it mixes with sample 18. Coriolis forces cause the diluent stream to pass along one side of loading cavity 7 and cuvette 8 in the manner illustrated. The diluent stream may be a washing liquid such as physiological saline or a reactant such as a solution of antibodies depending upon the particular analysis being performed and the stage of the analysis. Rapid and thorough mixing of sample and reactant or washing liquids, as achieved by the subject invention, has long been recognized as desirable and necessary for accurate analysis. Following addition of the diluent as illustrated in FIG. 5, particulates in the mixture are centrifugally sedimented against the outer wall 20 of cuvette 8 as shown in FIG. 6, leaving overlaying liquid 21. The
overlaying liquid 21 is then removed by applying vacuum through drainline 9 as shown in FIG. 7. Alternatively, the aperture 19 in plate 4 may be closed and air pending the particulates, and resedimenting. After washing, various reagents may be added and removed sequentially. Agglomeration may be measured by de termining the rate at which resuspended particles are sedimented using photometric techniques in the usual manner. By using a thin pencil of light such as obtained from a slit light source, the sedimentation of particles past a fixed radial position may be observed by observing the output of the photodetector. For measurement of sedimentation rate, two slit light sources spaced at different radial positions may be employed and the time required for a sedimenting boundary to pass from one radius to another measured.
Cuvette rotors made in accordance with the invention may be used for the purpose of sedimentation to clarify liquid containing a suspended particulate prior to determining absorbancy of the supernatant liquid remaining after sedimentation. Particulate phenomena, including agglomeration, lysis, fluorescence, and radioactivity, can be observed continuously by appropriate detecting means. Supernatant liquid phenomena, including optical density, light scattering, fluoresence, radioactivity, and light absorbancy, can also be observed.
The above description of one embodiment of the invention is offered for illustrative purposes and should not be interpreted in a limiting sense. For example, drainlines 9 may be reoriented to discharge into a vacuum chamber about the periphery of disk 5 or repositioned to communicate with the bottoms of respective cuvettes 8 so that washed cells or other particulates may be recovered from the cuvettes with the rotor assembly at rest for tests in other instruments. Such repositioning would require drainlines 9 to be inclined upward toward plate 4v to avoid cuvette drainage due to gravity alone while permitting vacuum or pressure forced recovery. Also, a wide variety and sequence of washing and reacting steps may be performed using the subject analyzer depending upon the particular analysis. It is intended, rather, that the invention be limited only by the scope of the claims appended hereto.
What is claimed is:
1. In a photometric analyzer including a rotor assembly defining a circular array of sample analysis cuvettes adapted to repeatedly pass between a light source and a photodetector for photometric analysis of the cuvette contents, the improved rotor assembly which defines:
a. a circular array of loading cavities disposed concentrically within said circular array of sample analysis cuvettes, each of said loading cuvettes being in radial alignment with a corresponding cuvette in said circular array of cuvettes;
b. sloped barriers interposed between said loading cavities and cuvettes to prevent transfer of liquid therebetween under static conditions while permitting centrifugal transfer of liquids from said loading cavities to corresponding cuvettes, said barriers each defining a radially oriented mixing passageway which is positioned angularly to one side of said barrier and extends through said barrier; and
c. a drain passageway communicating with each of said cuvettes at a point adjacent the radial extrem- 4. The improved rotor assembly of claim 1 wherein a central loading and distribution chamber is provided adjoining the radially inner edge of said loading cavities, and wherein said chamber has a serrated periphery for causing liquid fed therein to be substantially equally distributed to said loading cavities when said rotor assembly is rotating.