US 3135816 A
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G. L. RAM ETAL 3,135,816
COLLOIDAL DENSITOMETER 3 Shees--Sheei' l June 2, 1964 Filed Jan. 18, 1960 INVENTORS GERSON L. RAM ROBERT G. SALAMON A TORNEY June 2, 1964 G. L. RAM ETAL 3,135,816
coLLoInAL DENSITOMETER Filed Jan. 18, 1960 3 sheets-sheet 2 F l G. 2
mvENToRs j GERsoN 1 RAM ROBERT G. sALAMoN BY ,i
A TORNEY June 2, 1964 G. RAM ErAL' coLLoIDAL DENSITOMETER 3 Sheets-Sheet 3 Filed Jan. 18, 1960 FIG. 4
FIG/s INVENTORS GERSO N L. R A M ROBERT G. SALAMON ATTORNEY United States Patent O 3,135,816 CQLLOIDAL DENSITME'I'ER Gerson L. Bam, Passaic, and Robert G. Salamon, Bloomfield, NJ., assignors to Board of Trustees of Schools for industrial Education of Newark, New Jersey, Administering Newark College of Engineering, Newark, NJ., a public corporation of New Jersey Filed Jan. 18, 1960, Ser. No. 3,015 7 Claims. (Cl. 88-14) This invention relates to a device for measurement and precise recording of the degrees of opacity to the passage of light from a source of constant intensity through a scanned column of liquid, usually containing the matter being investigated as a suspension therein, so that the user may classify such matter and evaluate the nature, quality and characteristics thereof. The invention will enable such classification and evaluation to be attained for the iirst time in some cases and to a degree of accuracy and refinement in other cases not heretofore possible. For example, it will enable such matter as proteins, dissolved or suspended in said liquid, to be readily so evaluated and classified.
Probably the most unexplored, confused and chaotic eld of chemistry is that of the proteins. At the turn of the century, Abderhalden estimated that, on the basis of the amino acids known at his time, possible combinations of these acids would give 51/2 pentillion different protein compounds. It may be assumed on this basis alone, that the large number of compounds involved would make protein diterentiations extremely diilicult. In addition to the large number of proteins, another diliiculty stems from the fact that proteins are all very similar in chemical composition; for example, they all are compounds of hydrogen, oxygen and carbon containing about nitrogen. Consequently, the classical methods of analytical chemistry have given no simple system for the recognition of individual proteins. Such recognition of individual proteins is very important; in some cases being the diierence between life and death, since very closely related proteins such as albumens, coming from one animal species, are completely rejected when transferred into a second animal species. Yet, these diiferences cannot be determined or picked up by any simple chemical test, and at the present time, only the trial and error biological tests, such as in typing blood for transfusions, are generally known.
Another peculiarity which makes the differentiation of proteins a great problem is the capability of these substances or denaturation radically by such things as heat, light, radiation energy, or chemical agents. Again, this is a vital difference, representing the difference between life and death or health and disease.
Another factor involving the diiculties in working with proteins is the fact that these substances are capable of being used as food materials by various microorganisms and, consequently, will undergo microbial decomposition unless protected in a manner which does not denature the protein.
The present invention embodies the utilization of objective methods for the recognition of proteins. Ultimately, all chemical recognition problems are reducible to objective physical measurements. It makes possible the physical measurements of individual proteins suspended as colloids in a suitable liquid medium and protected from bacterial action, as by being held at a low temperature (approximately zero degrees centigrade). The physical measurements made by an instrument embodying this invention involve the tracking of the movements of protein colloids in a high voltage, extremely low-amperage direct current electrical iield. These protein colloids are suspended in a cold water solution of 3,135,816 Patented June 2, 1964 ICC non-ionic buffering character, desirably having a pH of about 8.6. The movement of the particles in the electrical field on a time basis is established by measuring light transmission across a column of liquid containing the protein under examination as a colloidal suspension, the entire length of the column of liquid being scanned by a moving light source. The small impediment odered by the particles to the transmission of light is greatly amplilied electronically by a photosensitive electron tube ampliiier mechanism.
Heretofore, optical systems of magnification were used to record changes in index of refraction. In the present invention electronic principles are used to measure impedance to direct transmission. Ultimately, several large categories of protein dilerentiation should emerge as experience in phyiscal measurement of the qualities described herein produces suricient data.
The disclosure herein of an illustrative use of the invention, for translation of the degree of opacity of matter in a column to passage of light from a source of constant intensity therethrough into visible indicia, is illustrative of a large number of other uses of the invention which will become apparent to those skilled in the art.
The drawings, illustrating procedures and devices useful in carrying out the invention, and the description below, are exemplary only of the invention, which shall be deemed to cover all other devices and procedures coming within the scope and purview of the appended claims.
In the drawings:
FIGURE 1 is a partly schematic side elevational view of apparatus embodying the invention,
FIGURE 2 is a fragmentary end elevational view thereof, taken at line 2-2 of FIGURE l, in the direction of the arrows,
FIGURE 3 is a fragmentary top plan view, with parts in horizontal section, taken at line 3-3 of FIGURE 2,
FIGURE 4 is a fragmentary elevational view of the cassette and its housing frame forming part of the apparatus of the preceding figures and embodying the invention, and
FIGURE 5 is a horizontal sectional view of the cassette and its frame on the line 5-5 of FIGURE 4.
Pursuant to the invention, a transparent or translucent container or cassette 10 (FIGURES 4 and 5) formed, for example, of a solid block of plastic or glass, is provided, and a pair of parallel chambers 11, 12 are cut therein and interconnected at their lower ends by a crossover passage or port 13 with a filling pipe 24, connecting therewith through a passage 23. At the upper end, passages, such as 14, 15 and 16, are formed connected through pipes or tubes 17 to the vessels 18 at the sides of the chambered block. Thus, the upper end of each chamber is connected to the adjacent vessel. The vessels 18 are interconnected thereabove by a bridge pipe 25 so that overow from one vessel will pass to the other. The chambers 11 and 12 may be vented, as by the pipes 19, 20 connected thereto by the passages 14. Cassette 10 may be suspended in a surrounding housing frame 22 as by a vertically adjustable rod 21. The vessels 18 are corked or otherwise closed as at 26, electrodes 27, 28 being projected into the vessels 18 and connected externally thereof with a power source (not shown) for imposing a unidirectional dierence in potential between said electrodes. The flow of current may be from electrode 27, through the liquid in tube 17 at left, passage 14, the column of liquid in chamber 11, passage 13, the column of liquid in chamber 12, passage 14, and tube 17 at right to electrode 28.
The housing frame 22 is sealed watertight as by and between a pair yof light-permeable plates 29, 29' (FIG- URES 1 and 5) the latter being also sealed to light hoods V39 from whence they emerge as a thin beam of light iti which may, if desired, be trimmed to a still smaller dimension, say about 1/16 in diameter, by passage through an aperture of corresponding size in a diaphragm (not shown). Such a beam 49 passes through the cassette 10 from the light source 33. The source 3S is movable vertically on the externally threaded shafts 4Z (FIGURE 2) journaled in the platform 43 for free rotation about vertical axes, said'shafts-also passing freely through bearings 44 on the front wall 37 of the tank (lGURE 1). Bevel gears d5 (FIGURES l and 2) are fixed to the upper ends of the vertical shafts l2 and mesh with bevel gears 46 of a shaft 47, which may be supported in bearings 52. A pinion 49 fixed on shaft 4'?, meshes with pinion 5t? of a second horizontal and drive shaft 51, passing freely through support bearings 52. The vertical shafts 42 may be rotated manually as by the hand wheel 63 (FlGURE 2,) keyed to shaft 51 or by the pulley 55 keyed to shaft 51 and drivenby abelt or other means Se actuated by a motor (not shown). A hand crank 53 may be keyed to one of the vertical shafts 4?. so that the shafts may thus be manually or automatically actuated as desired for a purpose now to be explained. The light source 3S may be secured to blocks 41 by a beam (nl (FlGURE 2) having slots 63 for the adjustable reception of threaded posts 64 fixed to bosses outstanding from the blocks 41 which are threaded on the shafts 4t2. Wing nuts 65 threaded on the posts 64, may be used to tighten the beam d@ at the desired position horizontally on the blocks 41.
Thus the light source 38 may be shifted to selective alignment with one of the chambers 11 and 12, or for realignment from one to the other rapidlyand accurately as desired. The blocks 41 are thus mounted for movement vertically on rotation of the shafts l2 to correspondingly move the light source 3S to act as a scanning device. The beam 66 may be provided with markings (not sh wn) for registering with markings (not shown) on the blocks 41, to precisely align the light source 33 with one of theVV chambers l1 and 12, as desired. The light source 3S should be constant in output and of sufficient intensity to generate a light beam 49 (FIGURES l and 3) of strength or intensity suiiicient for the purposes herein mentioned. A motorriven carriage is thus provided (beam 69, blocks 41, shafts 42 47, 51 and belt 56) so that the light source 38 may be moved at a constant or desired rate of speed verticallyV in line with a liquid in the selected one of the chambers 11 and 12 fortest or measurement of the degree of opacity of said liquid to passage of the light beam itl therethrough.
The housing frame 22 and associated parts are desirably immersed in a liquid in a tank 37, preferably refrigerated to about zero degrees C. To that end, a refrigerator or refrigerating urn't de (FlGURE l) may be provided with Yrefrigeration coils, fragments of which are shown schematically at 67, extended and disposed within the tank 34. Thek refrigeration 'prevents overheating of the solutions in the chambers 11, 12, by the electrical current owing therethrough, inhibits the growth of bacteria, and eliminates thermal currents which might upset boundary demarcations in the liquid or suspension in the chambers.
The 4power may be from any suitable current supply source to which electrodes 27, 2, are connected. A high voltage unidirectional source, not necessarily high powered, because only a small current llows, is desired, to produce a potential difference inducing electrophoretic operation. The apparatus of the invention electronically magniiies and measures the dierences in degree of optical density caused by the dierences in colloidal segregation along a cousiderble length of non-uniform solutions in one of the chambers 11 and 12, as the scanning device 3S tracks the length of the liquid. The particles measured are colloidal, the apparatus being used to measure the optical density of the materials which have no appreciableV tensity permanently recorded as a graph 59 where the l scattering effect and which have no significant resistance to the passage of light other than that the refractive index is slightly changed.
In operation, the colloidal matter, suspended in the test samples disposed in the chambers 11 and l2 of the cassette 1Q, is electrophoretically caused to migrate across a boundary formed with a buffering liquid, wll'le scanned by the light beam t. Said beam il has its rays directed substantially parallel to one another by the lens 39. Its source 38 is driven by a turning of thethreaded shafts 42. After passing through the cassette, the light beam 49 is picked up by a photoelectric cell 35 and the output inreadings can be calibrated and quantitatively compared with a solution of known characteristics. The cell 35 has a sensitive area large enough to receive the output beam of light at any position of its range from one chamber 11 to the other chamber 12 and over its entire range of Vertical movement.
In using the apparatus of the invention, the buffer (such as a cold water solution of Va non-ionic character, pH 8.6)V is rst poured through funnel 67 into the chambers 1.1 and 12 and then the test solution or suspension (which may be a protein) poured in, following the bufferV and moving up so that separation of the buffer and test solution occurs about halfway up the chambers 11 and 12. The bath in tank 34 is kept at constant refrigeration temperature and the apparatus is set up to track chamber 11 or chamber 12, ie., to vertically move the light source at a constant speed in line with said chamber. lf it is found that the migration of the colloidal suspension in the chamber so being tracked is in a reverse direction to the direction of vertical movement ofthe light source, the latter is shifted to the other chamber, to track the latter (wherein the migration of the colloidal suspension Will be reversed to that of the first tried chamber) and thus Will be suitable for the direction of tracking of the light source. The use of two chambers for the matter being tested is desirable as it is not always possible to predict the direction of migration or propagation. The possibility of the boundary or interface moving out of the eld of the light beam is thus obviated. The graph formed on the record of recorder 59 which is thus obtained, is calibrated against a graph of a standardized liquid.
The degree to which the beam 49 of Vlight is intercepted by the chamber 11 (or 12) and variations along the length of the chamber will correspondingly modulate the light beam. VIt will be noted from the disclosure herein that where a chart or graph is used to record the intensity of the residue of the light beam 40 after passage through the chamber 11 or 12 and on to the photoelectric cell 35 (any other light-current converter suitable for the use described may be used in lieu of the cell 35) the output current so generated passes through cable 36 in a Watertight tube 31 and through an amplifier 32 of desired characteristics and actuates a stylus 58 to make a record on a chart or yrecord sheet of recorder S9. Any other recorder may be used in lieu of'recorder 59. By thus recording the rate of migration, the material of which samples have been placed in the chambers 11 and 12 may be precisely evaluated and classiiied.
Having now described the invention in detail in accordance with the requirements of the patent statutes, those skilled in this art will have no diiliculty in making Ychanges and modifications in the individual parts or their relative assembly in order to meet specilic requirements or conditions. Such changes or modifications may be made Withoutrdeparting from the scope and spirit of the invention, as set forth in the following claims.
l. 1n a densitometer device for tracking the movement of particles suspended in a liquid column at a boundary with a buffer liquid, a light permeable container for said liquid column, means for impressing a difference in electrical potential across said boundary, a light source movably mounted relative to one face oi the container for directing a beam or" light through the liquid in the container, said light source being movable along the entire length of the container, a device beyond the other face of the container and sensitive to the light beam so passed through the column, means for effecting relative movement between the light source and the container at a constant rate of speed for scanning the full length ofthe contained liquid by said beam, and means for recording the intensity of the beam after passing through said column as a measure of the degree of opacity of said column throughout the entire length thereof to the passage of said beam as registered by said light sensitive device.
2. In a densitcuneterl device as set forth in claim 1, a light-permeable chamber in said light-permeable container for so suspending said particles in the liquid column, a supply pipe for lling said chamber, and air venting means for said chamber.
3. In a densitometer device as set forth in claim l, wherein the means for 'unpressing a difference in potential includes a vessel at each side of the container, an electrode in each vessel, and liquid passageways from said vessels to opposite ends of said column.
4. In a densitometer device as set forth in claim l, a second liquid column for alternative use disposed to one side of the rst-memtioned liquid column, and means connecting said rst and second liquid columns to allow liquid to flow therebetween.
5. in a densitometer device as set forth in claim 4, means for laterally moving said light source to remove the beam of light from one liquid column and selectively direct it to the other column.
6. In a densitometer device as set forth in claim 1,
wherein the container consists of a block of translucent material, a pair of chambers formed therein and parallel to one another, a passage in said material connecting the lower ends of said chambers, a lling pipe leading to said passage, a vessel disposed at each side of said block, and a passage in said block connecting the upper end of each chamber to the adjacent vessel.
7. In a densitorneter device as set forth in claim 1, wherein there is a housing frame surrounding and sealed watertight, to enclose said container, by means of lightpermeable plates, one in front and one behind said container, and means for adjustably suspending said container in said frame.
References Cited in the tile of this patent UNITED STATES PATENTS 2,430,895 Tuve et al. Nov. 18, 1947 2,495,297 Stern lan. 24, 1950 2,690,694 Wilson Oct. 5, 1954 2,816,479 Sloan Dec. 17, 1957 2,817,264 Pearson Dec. 24, 1957 2,834,247 Pickels May 13, 1958 2,843,003 Baker July 15, 1958 2,935,908 Phillips May l0, 1960 2,948,186 Kendall Aug. 9, 1960 FOREIGN PATENTS 938,937 Germany Feb. 9, 1956 OTHER REFERENCES Electrophoresis (Gray), published in Scientic Amer., volume 185, Issue 6, December 1951, pages 45-53.