US 3751357 A
Description (Le texte OCR peut contenir des erreurs.)
Aug. 7, 1973 Filed May 24, 1971 S. D. RAINS I ELECTROPHORESIS SYSTEM AND GEL FRAME 2 Sheets-Sheet 1 STEPHEN o. RAINS INVENTOR BERNARD D. BOGDON ATTORNEY Aug. 7, 1973 so. RAINS ,7
ELECTROPHORESIS SYSTEM AND GEL FRAME Filed May 24, 1971 2 Sheets-Sheet 2 32 v mzm l 1 I I I rll I ,7 [til I I", I I II I IIIIIIIIIIIIYIIIIIIIIlI/I I/II/IIIIIIIIIIIIIIIIIIJI FIG.3
f qi l l l l l l STEPHEN D. RAINS INVENTOR BERNARD D. BOGDON ATTORNEY United States Patent Office Patented Aug. 7, 1973 ABSTRACT OF THE DISCLOSURE BACKGROUND OF THE INVENTION 1) Cross-reference to related applications This invention is related to the invention described in the concurrently-filed and copending applications entitled, Method of and Apparatus for Depositing a Fluid in a Gel, Ser. No. 146,262, filed May 24, 1971, for inventors Stephen D. Rains and Leon L. Wheeless, .Tr., and Immunoelectrophoresis Gel Tray, Ser. No. 146,388, filed -May 24, 1971, for inventor Stephen D. Rains.
(2) Field of the invention This invention relates to apparatus and a system for carrying a gel for electrophoresis analysis of a fluid sample and, more particularly, to either self-contained or separate electrophoresis units for supporting a gel and solutions provided for connection to electrodes for energizing an electrical field across the gel having apparatus for introducing a fluid sample.
(3) Description of the prior art In the past, apparatus available for use in the field of electrophoresis has involved the use of awkward and cumbersome devices which frequently resulted in inaccurate and inconsistent results. For example, in the apparatus as disclosed in US. Pat. No. 3,432,414 entitled Electrophoretic Process With Continuous Scanning, issued to R. N. Rand on Mar. 11, 1969, there is disclosed, as best seen in FIG. 4, an apparatus for carrying a sample fluid comprising a string 49 stretched across an applicator 26 for positioning on the surface of the migration layer 6. It will readily be appreciated that there is a great deal I of difliculty experienced in consistentl depositing a fluid sample onto the string in the proper quantity without causing over-saturation to the string with the resulting blotting of the sample when the string is placed in position on the surface of the migration layer.
Another known method which is equally difficult for the hereinbefore mentioned processes involves the use of an applicator having the facility for introducing a fluid in a relatively thin line upon the gel. The disposition of the fluid upon the gel is controllable in thickness only. It is fairly obvious that the accuracy of the operator may vary as he draws the applicator across the gel which may, accordingly, result in a fluid deposition line of varying dimensions. Generally, this device has no means for controlling the speed of the movement of the operators hand when traversing the gel, the amount or rate of fluid being dispensed and further the direction of the motion. In addition, in the prior art devices it has generall been necessary to make the gel solution immediately prior to the test to be conducted and to cast the gel within the respective frames allowing it to set for a short period thereafter. The resulting multiplicity of steps complicated the job and created a number of unnecessary steps and further necessitated a great deal of the operators time to conduct each singular test.
Prior to applicants present invention the results obtained from the previously mentioned devices have been somewhat unsatisfactory and undependable. It was, at times, necessary in the past to repeat the tests to corrobrate and verify the previously obtained results.
SUMMARY OF THE INVENTION This invention provides for an electrophoresis gel frame having a migration portion interposed between, and in communication with, buffer chambers disposed at opposite ends of the frame for receiving electrodes to create an electrical field for causing migration of a fluid sample. The invention further provides for an accurate dependable device, easy and inexpensive to manufacture, convenient to store and handle and for all practical purposes disposable.
Embodiments according to the principles of the present invention can be constructed from a great variety of materials. It, however, has been found during the process of the experiments run that synthetic materials, particularly those belonging to the plastic family, work exceedingly well. The need for holding especially close tolerances which, correspondingly, increase manufacturing costs are essentially eliminated. This has decided advantages over the electrophoresis gel frames heretofore known in the field, particularly, for example, the gel frame disclosed in the previously mentioned R. N. Rand patent.
The present invention provides for including a gel in a migration portion of a gel frame in communication with buffer solutions contained in buffer chambers. The resulting gel frame may then be stored until a later date when the sample fluid will be introduced into the gel and the electrophoresis process carried out. This greatly facilitates set-up procedures, handling, and also reduces the time required for performing the electrophoresis operation. After completion of the operation for electrophoresis study, the operator may, if he so chooses, dispose of the frame without incurring great expense. The device of the present invention thereby further eliminates the need for a cleaning station and the accompanying materials necessary to prepare the frames for subsequent reuse. In addition, the device of the present invention will decrease the number of trained personnel necessary to perform the required operations.
A further embodiment of the present invention provides for an electrophoresis gel frame having separate buffer chambers, which for convenience purposes may or may not be fixed. A removable electrophoresis migration tray is provided to be bridged between the separate buifer chambers. The embodiment of this description may be constructed from a variety of materials, although, as previously discussed, synthetic materials, such as plastic, work very well.
In operation the buffer chambers have introduced into them the necessary substances to enable the electrophoresis process to take place. The prepared migration tray, or trays as the case may be, is then disposed so as to bridge the gap between the buffer chambers and also to make intimate contact with the gel in the tray and a compatible substance in the buffer chamber. A continuous path is thereby provided between the migration tray and the buffer chambers. Electrodes are disposed within the buffer chambers and then energized to facilitate the process of electrophoresis across the migration tray.
It is, of course, readily seen that the buffer chambers, as well as the migration tray, may be prepared in sepa rate and remote preparation areas, stored for a period of time and then brought together for use in the desired electrophoresis processes. It will also be easily seen that the buifer chambers may be used frequently and, particularly, may be used to conduct more than one test simultaneously. The resulting number of steps required to perform the operations, as well as the number of personnel required, will be favorably reduced.
BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a perspective view of an apparatus according to the principles of the present invention;
FIG. 2 is a top plan view of the apparatus;
FIG. 3 is a cross-sectional view of the apparatus of FIG. 2 taken along line 33 of FIG. 2; and
FIG. 4 is a perspective view of a further embodiment, according to the principles of the present invention, showing a plurality of buffer tanks having an electroplhoresis tray placed therebetween.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS In FIGS. 1, 2 and 3 a gel frame is shown which comprises a migration portion 12 extending between the bufier chambers 14 and 16 which are disposed at respective ends of the frame 10. The gel frame 10 generally shown in FIG. 1 consists of a migration portion 12 which comprises two spaced apart and parallelly disposed runners 18 and 20 extending between the buffer chambers 14 and 16 to define therebetween a generally rectangular aperture 22 for supporting a gel material 24 which is best seen in FIG. 2. The gel may comprise, for example, a medium such as is defined in US. Pat. No. 3,399,127 entitled Electrophoresis Medium Using Agarose and Carrageenan, issued to R. 'N. Rand et al.
A gelatinous fluid is made and then poured into the respective bufier chambers 14 and 16 through openings 15 and 17, respectively. The gelatinous fluid then proceeds to flow through openings 26 and 28, as best seen in FIG. 3, which communicate the buffer chambers 14 and 16 respectively with the rectangular aperture 22 of the migration portion 12 of the frame 10. The gelatinous fluid fills the rectangular aperture 22 completely while the buffer chambers 14 and 16 are filled to a level approximately three times the thickness of the fluid contained within the rectangular aperture 22. To provide for an even distribution of the gelatinous fluid within the rectangular aperture 22 of the migration portion 12 there may be provided a thin, relatively smooth and flat film or plate 30 positioned at the underside of the frame 10 for supporting the gel 24 in the fiuid state. A further thin, relatively smooth and flat film or plate 32 may be provided for containing the gelatinous fluid in the upper area of the migration portion 12, as best seen in FIG. 3. The use of thin films or plates 30 and 32 provide for uniformly regular and flat upper and lower surfaces on the gel 24. This feature will greatly insure accurate and true readings of the migration path across the migration portion 12. These readings may, of course, be made by any suitable known apparatus available in the field. It will be appreciated that any variations in the thickness of the gel would cause false and inaccurate readings when the migration path is analyzed and the use of the upper and lower thin films or plates 32, 30, respectively, greatly insures generally parallel upper and lower surfaces of the gel 24. An accurate reading is therefore assured. It will be further appreciated that the thin films or plates 30 and 32, respectively, would protect the gel during shipping, prevent dehydration during the process of electrophoresis and, further act as heat sinks and heat conduits.
A wire-like element 34, as best seen in FIG. 1, is provided for utilization in introducing a fluid sample into the formed gel 24. The wire-like element 34 extends through openings 36 and 38 in the runners 18 and 20 respectively of the migration portion 12. A fluid sample well 40 is positioned, for example, in migration runner 18 above and connected to the opening 36. As can easily be seen, the fluid sample may be disposed in the fluid well 40 by any convenient methods, such as, for example, those described in the hereinbefore concurrently-filled applications Ser. Nos. 146,262 and 146,388. Similarly, the fluid sample is disposed through the gel 24 by the wire-like element 34, as defined in the hereinbefore mentioned patent application.
After the gelatinous material is introduced into the frame 10, as previously described, and after it has set, a butter solution 19 is introduced through openings 15 and 17 of buffer chambers 14 and 16 respectively. The buffer solution 19 may be composed of any well-known buffer material such as those defined in the Rand Pat. No. 3,434,- 414. This arrangement, which is best seen in FIG. 2, provides for easy introduction of the electrophoresis electrodes without difiiculty in setting the apparatus up and without transferring the migration portion 12 from one work area to another. In addition, embodiments according to the present invention are such that they do not necessitate the need for battles such as those described in the immediately previously mentioned R. N. Rand patent. Batlies may, however, be incorporated to better control the electrophoresis process, if it is so desired.
After the bufler solution is introduced into the buffer chambers 14 and 16 end caps 42 and 44 may be placed over the openings 15 and 17 of the buffer chambers 14 and 16 respectively to thereby contain in cooperation with thin films 30 and 32 the gel and buffer constituents within the frame 10. The provision of the end caps 42 and 44 on the buffer chambers 14 and 1'6, respectively further provides for a convenient package for both handling and storage.
In an alternate embodiment illustrated in FIG. 4 there is shown an electrophoresis migration tray 50 bridged across two similarly constructed electrophoresis buffer tanks 52 and 54. The process of electrophoresis is similar to the process of electrophoresis hereinbefore discussed. The structures diifer, however, in that the tray 50 is, itself, devoid of buffer tanks. The migration tray 50 must therefore be placed upon the buffer tanks 52 and 54 to complete the electrophoresis process. After the electrophoresis process has been completed the migration tray 50- may be readily removed for analysis or replacement by a further migration tray. The embodiment of FIG. 4 is ideally suited for making the buffer tanks 52 and 54 as a relatively fixed structure for including an enclosure for performing the electrophoresis process. Alternately, the embodiments disclosed may be individually movable to enable an operator to selectively place the tanks as he desires. Additionally, it is easily appreciated that a plurality of trays 50 may be placed to bridge the area between the buffer tanks 52 and 54 in order to run any desired number of tests.
In the detail, the trays 50 may be similar to embodiments according to the principles of the invention included in the concurrently-filed Rains et al. patent application hereinbefore mentioned. The tray structure would be similar in that it would have a wire-like element 56 situated therethrough for introducing a fluid sample into a gel area 58 which is contained within and extends the length of the tray 50. The tray 50 is designed so that the gel area 58 is exposed at the bottom portion for placement upon the gel slabs 60 and 61 in each of the buffer tanks 52 and 54, respectively. The gel slabs 60, 61 are formed contiguous with one wall of the respective buflfer tanks 52 and 54 and extend slightly above the uppermost ledges 62 and 63 of the buffer tanks 52, 54, respectively, in order to make intimate contact with the bottom side 59 of the gel portion 58in the tray 50.
Butter solutions 64, 65 are provided within the buffer chambers 52 and 54 respectively so that intimate contact is made with the gel slabs 60 and 61. The buffer solutions 64 and 65 are further provided to receive therein electrodes 66 and 67, respectively, which may be formed, for example, of platinum wire. To preclude migration of the acid and base formed in an electrolytic process such as the one herein described, and more fully described in the Rand Pat. No. 3,432,414, baffie plates '68 and 69 are provided in buffer tanks 52 and 54, respectively, and positioned to be disposed between the electrodes 66, 67 and the gel slabs 60 and 61. The gel slabs 60 and 61 are formed in the baffle tanks 52 and 54 and about the baffle plates 68 and 69 so that a continuous path of buffer solution is provided around and under the baffle plates 68 and 69. The tray 50, the buffer tanks 52 and 54 and the bafile plates 68 and 69 may be formed of any suitable material, such as, for example, plastic. A power source 70 is provided which energizes the electrodes 66 and 67 and completes the process of electrophoresis.
1. A device for supporting a gel useable as a medium in analyzing chemical constituents 'by chemical separation, comprising:
a pair of receiving chambers containing buffer solution and a gel said chambers to be disposed apart from each other;
a generally flat and relatively thin frame having a perimeter defining an opening in the frame said frame positioned to communicate one chamber with the other chamber;
a gel disposed within and supported by the opening of the frame and formed to be in intimate contact with the gel in the chambers; and
means for providing a pressure reduction between a first point and a second point within the gel disposed within and supported by the opening in the frame to move chemical constituents from the first point to the second point within the gel.
2. The device as defined in claim 1, wherein the means is disposed within the gel and is a wire-like element.
3. The device as defined in claim 1, further comprising removable plates disposed upon the frame for positioning in intimate contact with the gel to be carried by the frame to assure smooth, parallel surfaces on the gel.
4. The device as defined in claim 1, further comprising covers for placement over the receiving chambers.
5. The device as described in claim 6, wherein the covers 'are removable.
6. The device as described in claim 7, wherein the covers are perforated to permit the insertion of electrodes therethrough and into the receiving chambers.
7. The device as defined in claim 1, further comprising means for introducting an electrical current to the chemical constituents to facilitate the electrophoresis process.
8. The device as defined in claim 7, wherein the means for introducing an electrical current to the chemical constituents are electrodes disposed within the buffer solution and extending the length thereof.
9. The device as defined in claim 8, wherein the electrodes are formed of a non-corrodible material.
10. The device as defined in claim 9, wherein the electrodes are formed of platinum.
11. The device as defined in claim 1, wherein the means for providing a pressure reduction between a first point and a second point within the gel is a filament to be disposed and movable within the supported gel from the first point to the second point to create a pressure reduction between said first and second points to move chemical constituents under the influence of the created pressure differential from the first point to the second point.
12. The device as defined in claim 2, wherein the wire like element is formed of a thread-like material.
13. A device for supporting a gel useable as a medium in analyping chemical constituents by chemical separation, comprising:
a pair of receiving chambers containing a buffer solution and gel said chambers to be spaced apart from each other;
a generally fiat and relatively thin frame having a perimeter defining an opening said frame positioned to communicate one receiving chamber with the other receiving chamber;
a gel disposed within and supported by the opening of the frame and formed to be in intimate contact with the gel in the receiving chambers; and
means for flowing chemical constituents by capillary action from a first point and a second point within the gel disposed within and supported by the generally flat and relatively thin frame.
14. The device as defined in claim 13 wherein the means for flowing chemical constituents from a first point to a second point within the gel is an element having absorbent qualities to permit the chemical constituents to be absorbed therein when the element is at least in part immersed in the chemical constituents.
15. The device as defined in claim 1, further comprising bafile plates disposed within the receiving chambers.
16. The device as described in claim 13, further comprising removable plates disposed upon the frame for positioning in intimate contact with the gel to be carried by the frame to assure smooth, parallel surfaces on the gel.
17. The device as defined in claim 13, further comprising removable covers for placement over the receiving chambers.
18. The device as defined in claim 13, further comprising means for introducing an electrical current to the chemical constituents to facilitate the electrophoresis process.
19. The device as defined in claim 18, wherein the means for introducing an electrical current to the chemical constituents are electrodes disposed within the buffer solution and extending the length thereof.
20. The device as defined in claim 13, further comprising baffle plates disposed within the receiving chambers.
References Cited UNITED STATES PATENTS 3,371,027 2/1968 La Paglia et al. 204-299 3,432,414 3/1969 Rand 204 G 3,470,080 9/1969 Raymond et al. 204180 G 3,479,265 11/1969 Elevitch 204180 G 3,554,894 1/ 1971 Zemel 204-180 G X 3,558,459 1/ 1971 Granstrand et al. 204180 G 3,616,387 10/1971 Siebert et a1. 204180 G 3,622,484 11/ 1971 Cawley 204180 G 3,674,678 7/1972 Post, Jr. et a1. 204-299 JOHN H. MACK, Primary Examiner A. C. PRESCOTT, Assistant Examiner