WO1997036170A1

WO1997036170A1 - Electrophoresis apparatus

Info

Publication number: WO1997036170A1
Application number: PCT/US1997/004934
Authority: WO
Inventors: Arthur H. Haeffner, Jr.; Michael W. Schuette
Original assignee: Life Technologies, Inc.
Priority date: 1996-03-27
Filing date: 1997-03-27
Publication date: 1997-10-02
Also published as: AU2548997A

Abstract

An electrophoresis apparatus (100). A first embodiment of the present invention is used for separating proteins or nucleic acids by performing electrophoresis. This embodiment includes a buffer tank (104) including therein an electrophoresis area. A first electrode is mounted on a first portion of the apparatus, and a second electrode is mounted on a second portion of the apparatus. A first conductor and a second conductor are disposed on the buffer tank (104), and provide conductive paths from a power source (120) to the first and second electrodes, respectively. A gel is placed in the buffer tank (104) and a pair of open-celled, foam pads saturated in a buffer solution are disposed in the buffer tank (104). A lid (136) is placed on top of the buffer tank.

Description

Electrophoresis Apparatus

Background ofthe Invention

Field ofthe Invention

The present invention relates to an apparatus for performing electrophoresis. In particular, the present invention relates to a low-cost electrophoresis apparatus.

Related Art

Electrophoresis is based on the principle that charged particles suspended between opposite poles in an electric field migrate toward the pole possessing the charge opposite that ofthe particle. The extent of migration is an indication of the composition of the particles. Electrophoretic separation is often used to separate DNA or RNA, referred to hereinafter as nucleic acid, or protein fragments generated as part of sequencing procedures.

A conventional apparatus for performing vertical gel electrophoresis is described in detail in U.S. Patent No. 4,773,984 to Flesher et al. incoφorated herein by reference. This apparatus includes a gel mold composed of two flat glass plates separated by thin strips placed at opposite edges. A polyacrylamide gel is cast between these plates. For horizontal gel electrophoresis, an agarose gel is used. The electrophoretic separation is carried out in this gel. The gel can be cast in a thin layer using a gel casting apparatus, such as the system disclosed in U.S. Patent No. 5,520,790 to Chopas et al, which is incoφorated herein by reference. Alternatively, a gel cast in a conventional gel cassette may be used. Wells are formed at one end ofthe gel for receiving samples to be tested. Typically, a micropipettor is used to place a small amount of a sample into each well.

Some apparatus are designed to orient the gel vertically; others are designed to orient the gel horizontally. A vertical gel electrophoresis apparatus typically includes a support platform having means for securing the gel to the support platform. This apparatus also has upper and lower reservoirs for containing a buffer solution. An electrode is installed in each reservoir to apply a voltage to the buffer solution. Placement ofthe gel against the support platform situates the gel so that when buffer solution is added to each of the reservoirs, an effective electrical contact is established between the buffer solution in one reservoir and the buffer solution in the other reservoir through the gel. Thus, a voltage differential is placed across the gel. For horizontal gel electrophoresis, the apparatus generally has one reservoir for holding a buffer solution. The gel is placed horizontally in the reservoir so that the buffer solution surrounds all sides ofthe gel. A thin layer of buffer solution usually covers the gel. Electrodes are disposed in the buffer solution on either side of the horizontal gel. Again, this places a voltage differential across the gel. This voltage difference causes protein or nucleic acids to separate from the samples into bands as they migrate along the length ofthe gel. The bands are then compared to one another for analysis.

Electrophoresis is generally performed using a relatively expensive electrophoresis apparatus and power source. Often, high voltages are used to perform electrophoresis of gels. The power source required to provide the high voltage is often costly. The cost factor involved in performing gel electrophoresis makes this process cost prohibitive for most schools and some research facilities. An inexpensive and convenient apparatus is needed for performing gel electrophoresis. Further, performing electrophoresis by manually loading the samples into wells in the gel, adding the buffer solution and running the samples through the gel, is a time consuming process. What is needed is an apparatus having buffer solution preloaded and capable of being placed through an automated system for loading the samples to be tested. Further, what is needed is an automated system that would load the samples into wells in the gel and apply the necessary electricity to cause electrophoretic separation ofthe samples through the gel. Still further, what is needed is an automated system that would photograph the bands in the gels after the electrophoresis process is complete for subsequent study and analysis.

Blotting is a method used in connection with gel electrophoresis to facilitate transfer ofthe bands on the gel to a transfer membrane. Once samples have been run through a gel and bands have formed, it is often desirable to transfer the bands from the gel to a membrane. The membrane can then be processed, using methods common to one skilled in the relevant art, to define proteins or nucleic acids on a nanometer scale. There are several methods typically used to transfer protein or nucleic acid bands from a gel to a membrane.

One method is through the use of capillary action. In the capillary method, a transfer membrane is placed on top of a gel. The gel is placed on a support tray which is disposed within a buffer solution reservoir. The support tray prevents the gel from directly touching the pool of buffer solution in the reservoir. A piece of filter paper is placed between the gel and the support tray. This filter paper drapes over the sides ofthe support tray and rests in the buffer solution. The filter paper absorbs a portion ofthe buffer solution so tl at the paper is saturated. A second piece of filter paper is placed on top of the transfer membrane. A blotter pad is then placed on top of this second piece of filter paper. The blotter pad absorbs the buffer solution from the pieces of filter paper. Thus, the buffer solution is drawn through the gel and the membrane and into the blotter pad. As the buffer solution travels up through the gel to the blotter pad located above the gel, the protein or nucleic acid bands slowly transfer from the gel onto the membrane. The transfer process, using the capillary method takes approximately 8 to 10 hours to complete. A second method used to transfer protein or nucleic acid bands onto a membrane is to apply a vacuum to the membrane side ofthe assembly described above. The vacuum decreases the amount of time necessary to perform the transfer process described above. However, the assembly required to perform this vacuum method is often costly. Further, the exact amount of vacuum applied to the apparatus must be carefully monitored, because too much vacuum destroys the transfer membrane.

A third method used to transfer protein or nucleic acid bands onto a membrane is commonly referred to as electroblotting. A semi-dry electroblotting technique includes the steps of placing the assembly described above in an electrophoretic area containing a thin layer of buffer solution, just enough to contact the edges of the blotter pads on either side of the gel. Then, a low voltage, high current power source is attached to the chamber, and electrophoresis is performed. The voltage difference running across the gel causes the protein or nucleic acid bands to be transferred to the membrane. A tank-style electroblotting process includes arranging the assembly of the gel, transfer membrane and filter paper as described above. The assembly has blotter pads on both sides. The entire assembly is then placed substantially vertically in a tank of buffer solution. The voltage difference running across the gel causes the protein or nucleic acid bands to be transferred to the membrane. However, as noted above, typical electrophoresis devices, including electroblotting devices, and the power sources needed to operate these devices are expensive. Thus, an apparatus is needed for inexpensively and conveniently performing electroblotting using an electrophoresis apparatus. Summary ofthe Invention

An electrophoresis apparatus for separating protein or nucleic acids from a sample and for transferring the protein or nucleic acids onto a membrane includes a buffer tank which may be a unitary member thermoformed from a plastic material. Because this apparatus may be made from relatively inexpensive plastic material, it is feasible to replace the entire apparatus cost effectively. The electrophoresis apparatus ofthe present invention may be constructed as a vertical gel electrophoresis apparatus, a horizontal gel electrophoresis apparatus or an electroblotting apparatus. In a first embodiment, a substantially horizontal gel electrophoresis apparatus is described. This apparatus includes a buffer tank which defines on a first side an electrophoresis area which is configured to contain a gel oriented substantially horizontally and a buffer solution. The buffer tank may also contain open-celled, foam pads disposed on either side ofthe gel that are capable of being saturated with buffer solution.

In the substantially horizontal gel electrophoresis apparatus, a first electrode is mounted on a first end of the electrophoresis area and a second electrode is mounted on a second end of the electrophoresis area. A first conductor is attached to a second side ofthe buffer tank to provide a conductive path from a power source to a point adjacent to the flrst electrode. A second conductor is similarly attached to a second side of the buffer tank to provide a second conductive path from the power source to a point adjacent to the second electrode.

The substantially horizontal gel electrophoresis apparatus also has a lid, preferably made from a transparent material, which is configured to mateably engage the buffer tank to enclose the electrophoresis area. A portion ofthe first conductor is disposed on the bottom side of the lid so that the first conductive path is completed when the lid is placed on top ofthe buffer tank. Similarly, a portion of the second conductor may also be disposed on the bottom side of the lid so that the second conductive path is completed when the lid is placed on top ofthe buffer tank. In a second embodiment, a substantially vertical gel electrophoresis apparatus is described. In the substantially vertical gel electrophoresis apparatus, a first electrode is mounted on a first end of the buffer tank and a second electrode is mounted on a lid. The lid is configured to mateably engage the buffer tank to enclose the electrophoresis area. A first conductor is attached to a second side of the buffer tank to provide a conductive path from a power source to a point adjacent to the first electrode. A second conductor is similarly attached to a second side of the buffer tank so that when the lid is disposed on top of the buffer tank a second conductive path is provided from the power source to a point adjacent to the second electrode. The substantially vertical electrophoresis apparatus include a tray for containing buffer solution. The tray is configured so that it may be disposed in the top ofthe buffer tank. Further the tray has at least one slot for receiving a gel mold assembly therethrough. In use, buffer solution is added to the bottom ofthe buffer tank and the gel mold assembly is inserted therein, in a substantially vertical orientation. Samples are then inserted into wells formed in the gel. The tray is then placed in the top of the buffer tank so that the top of the gel mold assembly extends updwardly through the slot in the tray. Buffer solution is then added to the tray. A power source, preferably a battery, is then connected to the apparatus so that electricity is delivered to the first and second electrodes to provide a voltage differential across the gel to perform electrophoresis.

In a third embodiment, an electroblotting apparatus is described in which the protein or nucleic acid is tranferred from the gel to a transfer membrane after electrophoretic separation of the sample has occurred. In a process commonly referred to as "electroblotting", the electroblotting apparatus is used to transfer the protein or nucleic acid bands on the gel to a transfer membrane. This transfer membrane can be made from nylon, nitrocellulose or polyvinyl difluoride (PVDF). The electroblotting apparatus similarly includes a buffer tank which can be formed from a material such as plastic and a first electrode mounted on one side of the buffer tank. The buffer tank is formed so that an electrophoresis area is defined therein. The first electrode is mounted at the bottom of this area.

A lid having a similar configuration to the buffer tank is also formed from a material, preferably plastic. This lid has a second electrode mounted on the bottom side ofthe lid. A protein or nucleic acid transfer package is placed in the electrophoresis area in the buffer tank. This transfer package includes first and second blotter pads with the gel and a transfer membrane sandwiched between the blotter pads. Filters are disposed between one blotter pad and the gel and the other blotter pad and the transfer membrane.

A user soaks the blotter pads, filters and transfer membrane in buffer solution and places the transfer package in the buffer tank. Alternatively, the blotter pads may be made from an open-celled foam so that the transfer package is presoaked with buffer solution. In this case, the user does not soak the transfer package. Then the lid is snugly fit on top of the buffer tank. In the preferred embodiment, the lid and buffer tank have tabs formed therein. The tabs are configured to mate with an adaptor. The adaptor is configured to be connected to a remote low-voltage, high-current power source. When the adaptor is secured on the tabs of the apparatus and the remote power source is connected to the adaptor, a conductive path between the first and second electrodes of the apparatus is provided. The electroblotting process takes between an hour and an hour and a half to complete the transfer ofthe protein or nucleic acid from the gel to the transfer membrane. After the protein or nucleic acid has been transferred, the bands are disposed on the transfer membrane. The transfer membrane can be further processed to identify the protein or nucleic acid bands on the membrane. Electroblotting allows a user to detect protein or nucleic acid on a nanometer scale. Thus, this electroblotting apparatus is also a less expensive alternative to a conventional electroblotting device. The present invention would allow researchers, who normally use the capillary method for protein or nucleic acid transfer due to its low cost, to use the faster and more efficient method of electroblotting.

In all three embodiments discussed herein, the first and second electrodes and first and second conductors may be made from a nickel foil having an adhesive on one side so that the electrodes and the conductors may be securely attached to the buffer tank and the lid. When the electrodes corrode and become unusable after several runs, either the electrodes or the entire apparatus may be replaced.

Further, in all three embodiments, the buffer tank may be molded from a plastic or other material to form a recess on the second side for receiving a power source, preferably a battery. The buffer tank may also include a registration key molded on a second side thereof for orienting the position ofthe wells in the gel with respect to the buffer tank ofthe apparatus. This registration key allows for samples to be loaded in the wells ofthe gel using an automated system.

These electrophoresis apparatus ofthe present invention may be sold as ready-to-use packages. For example, the substantially vertical and substantially horizontal gel electrophoresis apparatus may be sold as a package including a molded buffer tank and a lid, two electrodes, one or more conductors, at least one precast gel, a buffer solution either separate from the package to be added later or suspended in open-celled, foam pads, and a power source. The electrophoresis area in the buffer tank is sealed by placing the lid on top of the buffer tank.

Further, the lid may be heat sealed to ensure that there is no loss of buffer solution during shipping.

Because the package includes a precast gel, the user does not have to cast his own gel, which includes either mixing and heating agarose and applying the agarose to a gel slab for casting or casting a polyacrylamide gel. This time- consuming process is avoided by providing a precast gel with the electrophoresis apparatus.

To manually use the ready-to-use electrophoresis kit, a user simply breaks the heat seal to remove the lid, and loads the samples for testing into wells foimed in the precast gel. The user then places the buffer solution in the buffer tank(s) so that it surrounds the gel. If the apparatus was shipped with the foam pads saturated with buffer solution, this step is omitted. The packaged apparatus may be powered by a power source, for example, a 9-volt battery which is inserted into the recess on the second side of the buffer tank. The user simply places the lid on top ofthe buffer tank securely so that a conductive path is completed and electrophoresis is performed.

Further, in the case of the electroblotting apparatus, the ready-to-use package includes a molded buffer tank having an electrode mounted thereon, a lid having another electrode mounted thereon, and a protein or nucleic acid transfer package. Thus, in use, the user opens the ready-to-use electroblotting kit, inserts the gel into the transfer package, adds buffer solution and applies a power source to cause electrophoretic transfer ofthe protein or nucleic from the gel to the transfer membrane. These electrophoresis apparatus are conducive for school or research use because they are less expensive and safer than conventional electrophoresis apparatus. Because the present invention uses a smaller power source than a conventional electrophoresis apparatus, there is less risk of accidental injury of a user from electrocution. The substantially vertical and substantially horizontal electrophoresis apparatus could also be loaded and run in an automated system. The registration key is used by the automated system to locate the wells in the gel relative to the apparatus. An automated loader having micropipette tips automatically loads the samples to be tested into the wells. In one system, a device can be used to puncture holes in the lid through which the samples may be loaded. In another system, the lid is removed before loading the samples in the wells. The automated system then applies the necessary electricity to run the electrophoresis process. The automated system may also photograph the results ofthe testing for subsequent study and analysis.

Thus, the present invention provides the user with an apparatus for inexpensively conducting electrophoresis both to separate a protein or nucleic acid from a sample and to transfer protein or nucleic acids from a gel to a transfer membrane for further analysis.

Brief Description of the Figures

The foregoing and other features and advantages ofthe invention will be apparent from the following, more particular description of a preferred embodiment ofthe invention, as illustrated in the accompanying drawings.

FIG. 1 is an exploded view a substantially horizontal gel electrophoresis apparatus ofthe present invention having a buffer tank and a lid.

FIG. 2 is a perspective view ofa second side of a buffer tank of FIG. 1. FIG. 3 is a perspective view ofthe second side ofthe buffer tank of FIG. 1.

FIG. 4 is a top view of a first side ofthe buffer tank of FIG. 1. FIG. 5 is a simplified perspective view ofthe first side ofthe buffer tank of FIG. 1.

FIG. 6 is an end view ofthe buffer tank ofthe buffer tank of FIG. 4. FIG. 7 is a sectional side view ofthe buffer tank ofthe present invention taken along line A-A of FIG. 4. ^' FIG. 8 is a top view of a lid of FIG. 1. FIG. 9 is a bottom view ofthe lid of FIG. 1.

FIG. 10 is a top view of an altemate embodiment of the buffer tank of FIG. 4.

FIG. 1 1 is an exploded view of a second embodiment of the electrophoresis apparatus ofthe present invention.

FIG. 12 is an exploded view ofa substantially vertical gel electrophoresis apparatus ofthe present invention having a buffer tank and a lid.

FIG. 13 is a bottom view ofthe lid ofthe apparatus of FIG. 12.

FIG. 14 is a bottom view ofa second side ofthe buffer tank of FIG. 12. FIG. 15 is an end view ofthe buffer tank of FIG. 12.

FIG. 16 is a perspective view of an adaptor and a remote power source.

FIG. 17 is a sectional side view ofthe adaptor of FIG. 16.

FIG. 18 is a sectional side view of the adaptor of FIG. 16 matingly engaged with the apparatus of FIG. 11. FIG. 19 is a perspective view of an altemate embodiment of an adaptor matingly engaged with the apparatus of FIG. 11.

FIG. 20 is a top perspective view of an altemate embodiment of a lid of the apparatus of FIG. 11.

FIG. 21 is a top perspective view of an altemate embodiment ofa buffer tank ofthe apparatus of FIG. 1 1.

FIG.22 is a sectional side view of an alternate embodiment ofa horizontal gel electrophoresis apparatus ofthe present invention having a buffer tank and a lid.

Detailed Description ofthe Preferred Embodiments

A preferred embodiment ofthe present invention is now described with reference to the figures where like reference numbers indicate identical or functionally similar elements. Also in the figures, the left most digit of each reference number corresponds to the figure in which the reference number is first used. While specific configurations and arrangements are discussed, it should be understood that this is done for illustrative puφoses only. A person skilled in the relevant art will recognize that other configurations and arrangements can be used without departing from the spirit and scope ofthe invention. It will be apparent to a person skilled in the relevant art that this invention can also be employed in a variety of other devices and applications.

FIG. 1 shows an exploded view of substantially horizontal gel electrophoresis apparatus 100 ofthe present invention. Apparatus 100 includes a buffer tank 104, and a Hd 108. In all three devices discussed herein (i.e., a substantially horizontal electrophoresis apparatus, a substantially vertical electrophoresis apparatus, and an electroblotting apparatus), the buffer tank is preferably thermoformed from one ofthe following: polyvinyl chloride (PVC), phenylethyl thiogalactoside (PETG), or polystyrene. However, the buffer tank may also be injection-molded or otherwise formed from any plastic or other similar material, as would be apparent to one skilled in the art. Further, the buffer tank is preferably constructed as a unitary piece of the electrophoresis apparatus.

Buffer tank 104 has a lip 112 and a recess 116 for housing a power source 120. In all three embodiments discussed herein, a battery could be used as the power source. A conventional nine-volt transistor battery is one example of a power source that may be used in the electrophoresis apparatus. However, it would be apparent to one skilled in the relevant art that many types and sizes of batteries could be used. For example, a rechargeable battery could be used. Altematively, the power source could be an adaptor that can be plugged on one end into the apparatus and on the other end into a conventional wall socket. An example of such an adaptor is shown in FIGs. 16-18 and will be discussed in further detail below. The adaptor converts AC input to DC input for performing electrophoresis.

In all three devices, a recess may be formed in the apparatus for receiving a battery therein. For example, as shown in FIG. 1, a recess 116 is molded in buffer tank 104 so that a nine-volt battery is snugly secured therein. It would be apparent to one skilled in the relevant art that the recess could also be formed on one of the sides ofthe buffer tank or in the lid ofthe electrophoresis apparatus.

Further, the power source could be independent ofthe apparatus and connected via banana clips or similar connector devices to the electrodes ofthe apparatus.

FIG. 22 shows a sectional side view of an altemate embodiment of a substantially horizontal electrophoresis apparatus 2200. In the embodiment shown in FIG. 22, a registration key 2210 is molded in buffer tank 104 adjacent recess 116. The registration key is shown within the context of a horizontal electrophoresis apparatus by way of example only. It would be apparent to one skilled in the relevant art, that a similar registration key could be molded into a vertical electrophoresis apparatus.

Registration key 2210 is a distinctively-shaped notch or trough molded in buffer tank 104 and serves as an index for an automated system for locating wells 2208 in gel 2206 disposed in buffer tank 104. Gel 2206 includes a conventional gel mold assembly consisting of a two plates and spacers placed therebetween. The first and second plates are brought together in a planar fashion with spacers therebetween to form a gap the same thickness as the spacers. A gel slab is molded in gap so that the desired electrophoretic action can occur therein. A comb is typically placed in one end of the gel slab before the slab has completely solidified. The comb creates wells in the gel for housing samples for testing. The comb is removed, after the gel has polymerized. The gel mold assembly may be a conventional gel mold cassette or a conventional gel mold assembly including glass plates and spacer gaskets therebetween.

Gel 2206 is disposed in buffer tank 104 so that wells 2208 are in a known relationship to buffer tank 104. Once wells 2208 are located, an automated loader

(not shown) loads samples into the wells. Registration key 2210 must be shaped so that the automated system can correctly orient itself with respect to buffer tank 104 and gel 2206 disposed therein. The shape of wells 2208 can be formed by using a variety of comb-shapes to take into account minor tolerance errors. For example, in one embodiment, wells 2208 could be D-shaped so that the micropipettor tips on the automatic loader can easily locate wells 2208 and load the samples. Wells 2208 are more rectangular than a conventional well to accommodate the tips of the automatic loader and to minimize the amount of accuracy needed to load the samples. Buffer tank 104 has a first side 128, which is shown in further detail in

FIGs. 4 and 5. Indentations 132 are thermoformed into second side 124 of buffer tank 104. Indentations 132 provide feet for buffer tank 104 so that the buffer tank is level when resting on a working surface. In an altemate embodiment, one or more of indentations 132 could be formed so as to be used as a registration key in an automated system for locating wells 2208 disposed within buffer tank 104.

However, in a preferred embodiment, registration key 2210 is located as close to gel 2206 as possible to minimize the build-up of tolerance errors.

As shown in FIG. 1, Hd 108 has a projection 136 that is configured to mateably engage with lip 112 of buffer tank 104, so that when lid 108 is placed on top of buffer tank 104, projection 136 fits snugly inside first side 128 of buffer tank 104.

Buffer tank 104 is molded to form an electrophoresis area 420 (as shown in FIG. 4) to contain a gel and a buffer solution. In use, gel 2206 (shown in FIG. 22) is secured within electrophoresis area 420 so that it rests horizontally with respect to the working surface. For ease of explanation, the example apparatus shown in FIG. 22, shows only one gel disposed therein. However, in all three devices discussed herein, any number of gels could be disposed in each apparatus for performing electrophoresis. For example, substantially horizontal electrophoresis apparatus 100 and 2200 could be configured so that several gels could be lined up side-by-side in a substantially horizontal orientation between the first and second electrodes. Similarly, in a substantially vertical electrophoresis apparatus, described in further detail below, several gels could be stacked plate-to-plate in a substantially vertical orientation in the buffer tank. Still further, in the electroblotting apparatus discussed in further detail below, the apparatus could be molded to contain several transfer packages, each transfer package containing, among other things, a gel.

As shown in FIG. 22, gel 2206 rests on the top of recess 116, and open- celled, foam pads 2202 and 2204 are disposed in buffer tank 104 on either side of gel 2206. It would be apparent to one skilled in the relevant art that open- celled, foam pads could be similarly used in a vertical electrophoresis apparatus and in an electroblotter. Such uses of foam pads will be discussed in further detail below. Buffer solution is added such that it barely covers the top of gel 2206. Foam pads 2202 and 2204 fill the volume formed in electrophoresis area 420 of buffer tank 104 so that they reduce the amount of buffer solution needed to fill the tank. Further, foam pads 2202 and 2204 reduce the amount of evaporation of the buffer solution, because they reduce the surface area of the exposed buffer solution in buffer tank 104. Finally, foam pads 2202 and 2204 act also a contact spring with conductors 204 and 208 disposed on lid 108 so that it is easier to make electrical contact between power source 120 and gel 2206.

The buffer solution is added to buffer tank 104 to cover gel 2206 to prevent the gel from drying and to control pH of the materials inside the electrophoresis area. Further, the buffer solution cools gel 2206, which may otherwise retain heat generated during the electrophoresis process. The buffer solution is also slightly conductive, so that electricity from power source 120 is transferred across gel 2206.

In use, a user loads samples into wells 2208 which have been formed in gel 2206 during casting ofthe gel. A micropipettor (not shown) is generally used to deposit a small amount of each sample in well 2208 of gel 2206. In one embodiment, the apparatus is sent to the user with a precast gel. The gel may be cast using a conventional gel casting cassette, discussed above.

Altematively, using the embodiment shown in FIG. 22, gel 2206 may be cast directly into buffer tank 104 by pouring the molten gel into the area formed by the top of recess 116 of buffer tank 104 and the edges of foam pads 2202 and

2204. The density of the open-celled pores of foam pads 2202 and 2204 may be varied so that they absorb little, if any, ofthe molten gel. A comb is then placed at one end ofthe gel to form wells 2208, and gel 2206 is allowed to solidify. The comb is removed before running the apparatus, and samples are loaded into wells 2208.

A conductive path is completed between power source 120 and gel 2206 by the buffer solution. This conductive path will be described in further detail below. The electricity passing across gel 2206 causes the protein or nucleic acid in each sample to separate into bands as the sample migrates across the gel. The resulting bands are then compared for subsequent analysis and diagnosis.

Referring now to FIGs. 2 and 3, a perspective view of buffer tank 104 is shown. Second side 124 of buffer tank 104 has a first conductor 204 and a second conductor 208 disposed thereon. In all three devices discussed herein, the conductors can be made from any electrically conductive material. In the preferred embodiment, the conductors are made from a nickel tape, similar to

Flectron tape, available from Stockwell Rubber Company, Philadelphia, PA, as item number 3027-217. The nickel tape contains an adhesive on one side which allows the tape to be securely fastened to the buffer tank In another embodiment, a stainless steel tape can be used, available from Compac Industries, Inc., Edison, NJ. The stainless steel tape is capable of undergoing more runs of the electrophoresis process before corroding and becoming useless. The nickel tape has a tendency to corrode faster than the stainless tape, thus requiring more frequent replacement. However, the nickel tape is generally less expensive than the stainless steel tape.

As is apparent to one skilled in the relevant art, other conductive materials may be used for the conductors in all three devices. For example, carbon fiber could be used as a conductor. However, it has been noted that carbon fiber has absorbent qualities which cause the carbon fiber material to absorb the buffer solution in the buffer tank. Thus, the carbon fiber wicks away the buffer solution to the outside of the buffer tank Thus, this material is not preferable for the present invention. Other materials, such as thin-film metal tapes or conductive polymers, can be used as conductors. Further, metal materials can be deposited onto the surface ofthe apparatus to form conductors. In one embodiment, metal materials are deposited by vapor deposition or by plasma spray on to the surface ofthe apparatus.

First conductor 204 is disposed along a second side 124 of buffer tank 104 to provide a first conductive path from power source 120 to a first electrode 404

(as shown in FIG. 4). A portion of first conductor 204 is wrapped around lip 112 so that it is secured to first side 128 of buffer tank 104 (as shown in FIGs. 4 and

22).

Referring now to FIG. 4, a top view of first side 128 ofbuffer tank 104 is shown. As described above, first conductor 204 and second conductor 208 are shown on the first side of lip 112. First electrode 404 is shown adjacent first conductor 204 on lip 1 12. First electrode 404 extends downwardly into an electrophoretic area 420 created inside buffer tank 104. Similarly, a second electrode 408 is shown adjacent second conductor 208 on lip 112. Second electrode 408 extends downwardly into electrophoretic area 420. Electrodes 404 and 408 extend downwardly all the way to the bottom of buffer tank 104 so that when buffer solution is poured into buffer tank 104, a portion of electrodes 404 and 408 come into contact with the buffer solution.

In all three devices discussed herein, the electrodes can be made from Nickel tape. Altematively, the electrodes could be made from stainless steel tape. As discussed above with respect to the conductors, other metal foil materials can be used to make the electrodes, as would be apparent to one of skilled in the relevant art. Similarly, as discussed above, metal materials can be deposited onto the buffer tank to form the electrodes ofthe present invention.

An upward projection of recess 116 is shown on first side 128 of buffer tank 104. A gel is placed horizontally on top of and secured to this projection of recess 116. Also shown in FIG. 4 is a recess 412 formed in lip 112. This recess provides a means for the user to easily remove lid 108 from buffer tank 104 for disassembly of apparatus 100. During assembly of any of the three devices described herein, the lid may be heat sealed to the buffer tank using a conventional heat sealing process. This heat sealing process allows each apparatus to be shipped with the buffer solution already disposed in the buffer tank Thus, the user receives the apparatus virtually ready-to-use. Further, a removable foil piece may be placed over the top ofthe buffer tank prior to heat sealing the lid on top. This foil piece will prevent evaporation of the buffer solution during shipping and can be easily removed prior to running the apparatus.

FIG. 5 is a simplified perspective view of buffer tank 104 showing electrophoretic area 420. As shown in FIG. 5, first electrode 404 is an L-shaped piece of material. A small portion of first conductor 204 is also visible on lip 112. Although second electrode 408 is not visible from this perspective view, in the preferred embodiment, the shape of second electrode 408 matches that shown of first electrode 404.

Referring now to FIGs. 6 and 7, an end view and a sectional side view of buffer tank 104 are shown. First conductor 204 extends along second side 124 of buffer tank 104 and onto first side 128. First electrode 404 is shown by the dashed line on buffer tank 104. FIG. 7 shows buffer tank 104 having recess 116 for housing power source 120.

Referring now to FIGs. 8 and 9, a top and bottom view respectively of lid 108 are shown. As shown in FIG. 8, projection 136 extends downwardly from the top side of lid 108 to mate with buffer tank 104. A portion of first conductor 204 and second conductor 208 are disposed on the bottom side of lid 108. Thus, when lid 108 is placed on top of buffer tank 104, a portion of first conductor 204 electrically connects the remaining portion of first conductor 204 on Hp 112 with first electrode 404. Similarly, a portion of second conductor 208 electrically connects the remaining portion of second conductor 208 on lip 112 with second electrode 408. This configuration insures the safety ofthe user, because when lid 108 is removed from buffer tank 104, the electrical connection is broken, thereby considerably lowering any risk ofthe user receiving an electric shock. FIG. 10 shows an altemate embodiment ofthe present invention in which both conductors 204 and 208 on lip 1 12 are electrically connected to electrodes 404 and 408 such that an electrical connection is completed as soon as power source 120 is secured in recess 1 16. In another embodiment, only one of conductors 204 and 208 could be electrically connected to electrodes 404 and 408. In this embodiment, a portion of the conductor that is not electrically connected to the electrode, is disposed on lid 108. Thus, when lid 108 is secured on top of buffer tank 104, the portion of the conductor on lid 108 would electrically connect the remaining portion ofthe conductor with the electrode on only one side of lip 112. In the preferred embodiment, lid 108 is transparent so that the user may observe the protein or nucleic acid separating along the gel during electrophoresis. In an automated system, once the electrophoretic separation is complete, the system photographs the resultant bands in the gel for subsequent study and analysis. FIG. 11 shows an electrophoresis apparatus 1 100 which can be used to perform electroblotting. For example, apparatus 1100 can be used to transfer protein or nucleic acids bands from a gel to a transfer membrane, after apparatus

100 has been used to perform electrophoresis on the gel to cause the proteins or nucleic acids to migrate to form the bands on the gel.

Apparatus 1100 includes a buffer tank 1104 and a Hd 1108. Apparatus 1100 also includes a protein or nucleic acid transfer package 1112, hereinafter referred to as transfer package 1112. Buffer tank 1104 has a first electrode 1116 mounted thereon. A second electrode 1120 is mounted on the bottom side of lid 1108. It was discovered that at the higher currents needed to perform electroblotting, the Nickel on the electrodes deplated from the tape backing during the electroblotting process. Thus, in the preferred embodiment, the stainless steel tape described above is used for first and second electrodes 1116 and 1120. Buffer tank 1104 defines therein an electrophoresis area 1124. Apparatus

1100 is used after separating protein or nucleic acid on a gel for transferring the protein or nucleic acid on the gel to a transfer membrane. This allows a user to process the transfer membrane subsequent to the transfer, to make visible protein or nucleic acids on a nanometer scale. Thus, this electroblotting technique allows very small proteins or nucleic acids to be identified after separation.

Transfer package 1112 includes a first blotter pad 1128 and a second blotter pad 1144. A gel 1140 and a transfer membrane 1136 are sandwiched between first and second blotter pads 1128 and 1144, as shown in FIG. 11. The transfer package 1112 also includes a first filter 1132 located between first blotter pad 1128 and transfer membrane 1136, and a second filter 1 142 located between second blotter pad 1144 and gel 1 140. In the preferred embodiment, first and second filters 1132 and 1142 each consist of two sheets of 3 mm filter paper. First and second blotter pads 1128, 1144, first and second filters 1132, 1142 and transfer membrane 1136 are all soaked in buffer solution before performing the electroblotting. In one embodiment, first and second blotter pads 1128, 1144 are made from open-cell, foam pads. Transfer package 1 112 is placed within electrophoresis area 1124, and lid 1 108 is snugly placed on top of buffer tank 1104. In the embodiment shown, notches 1152 on buffer tank 1104 and lid 1108 form a tab 1146 on which first electrode 1116 is disposed and a similar tab 1148 on which second electrode 1120 is disposed, respectively. These tabs are flexible so that they can bend up and down slightly in response to an external force. As shown on lid 1108, second electrode 1120 is wrapped around the end of tab 1148 so that it is disposed on both sides of the tab. Similarly, first electrode 1 116 is wrapped around the end of tab 1146. These tabs 1146 and 1148 are connected to an adaptor (shown in FIG. 16) which is connected to an external power source to provide electricity for performing the electrophoresis process. First electrode 1116 acts as a cathode and second electrode 1120 acts as a diode so that the voltage differential causes the protein or nucleic acid to travel downwardly onto transfer membrane 1136. In an altemate embodiment (not shown), the electroblotting apparatus could be configured, as described above with respect to apparatus 100 and 2200, to house a battery as a power source.

In the preferred embodiment, when lid 1108 and buffer tank 1104 are matingly combined, an adaptor 1600, shown in FIG. 16, is placed onto tabs 1146 and 1148 to provide a conductive path between first and second electrodes 1116 and 1120. Adaptor 1600 provides a means for connecting an adaptor pin 1616 of a remote power source 1604 to apparatus 1 100 without exposing electrodes 1116, 1120 to the user. Because, electroblotting apparatus 1100 and electrophoresis apparatus 100 and 1200 (described below) may be operated using a power source having a high voltage, thereby posing a risk of danger to a user, an adaptor may be used to lower the risk of accidental electrocution during use. Adaptor 1600 consists of a wedge 1608 molded from a plastic material and housed in an enclosure 1612. FIG. 17 shows a sectional side view of adaptor 1600. Adaptor 1600 has a hole 1704 formed therein for receiving adaptor pin 1616. Adaptor pin 1616 is press fit into hole 1704 to hook up power source 1604 to apparatus 1100. Wedge 1608 is formed on the inside of enclosure 1612. Power source 1604 used in the preferred embodiment to perform electrophoresis provides a low voltage, high current power source. In the preferred embodiment, the power source provides a voltage in the range of 15 V to 18 V, and a current up to 1 Ampere. In an altemate embodiment, the voltage used may be in the range of 15 V to 25 V. Further, apparatus 1100 is constructed to be capable of undergoing electroblotting using a voltage as high as 150 V.

Adaptor 1600 has a first tab receiving area 1620 and a second tab receiving area 1624 formed on either side of wedge 1608. As shown in FIGs. 17 and 18, tab 1146 is shown disposed within tab receiving area 1620 and tab 1148 is shown disposed within tab receiving area 1624. As the wedge moves onto the tabs, first and second foil electrodes 1116 and 1120 are pressed against a first conductor 1708 and a second conductor 1712, respectively.

First and second conductors 1708 and 1712 are mounted on the inner side walls of enclosure 1612 and wrap around the inside of adaptor 1600 to also line the inner walls of hole 1704. First and second conductors 1708 and 1712 are preferably made from a foil. However, conductors 1708 and 1712 could be made from other materials as discussed above. In the preferred embodiment, first and second conductors 1708 and 1712 are made from stainless steel foil tape having an adhesive on one side so that the conductors can be secured to the inner walls of adaptor 1600. Referring now to FIGs. 19 - 21, an altemate embodiment of an adaptor

1900 is shown for use with any ofthe apparatus discussed herein. However, for ease of discussion, adaptor 1900 will be shown matingly engaged with apparatus 1100. Adaptor 1900 functions in the same manner as adaptor 1600, except that adaptor 1900 is key-shaped so that the user cannot accidentally reverse the connection. Thus, adaptor 1900 allows-the user to insert the adaptor on only one direction. Adaptor 1900 is configured to mate with adaptor pin 1616. FIGs. 20 and 21 show altemate configurations for lid 1108 and buffer tank 1104, respectively, so that apparatus 1100 can matingly engage adaptor 1900. A top perspective view of lid 1108 is shown in FIG. 20. Lid 1108 includes second electrode 1120 (shown with dashed lines) disposed on the bottom side ofthe lid. Tab 1148 is formed on the periphery of lid 1108 by notches 1152. As shown, second electrode 1120 wraps around the edge of tab 1148 and on the top of tab 1148. A top perspective view of an altemate embodiment of buffer tank 1 104 is shown in FIG. 21. In tiiis embodiment, a recess 2104 is cut away from the side of buffer tank 1104 to receive adaptor 1900. As shown, first electrode 1116 is secured to the bottom of buffer tank 1104 and is wrapped around the edge of tab 1144 formed on the periphery of buffer tank 1 104. FIG. 12 is an exploded view ofa substantially vertical apparatus 1200 of the present invention having a buffer tank 1204 and a lid 1208. In this embodiment, apparatus 1200 may be thermoformed from a plastic, as described above. Apparatus 1200 is molded to form therein an electrophoresis area 1212. Apparatus 1200 shows two gels 1220 disposed substantially vertically in electrophoresis area 1212. However, as discussed above, any number of gels could be stacked in buffer tank 1204.

Buffer tank 1204 has a first side 1224 and a second side 1228. A first conductor 1504 (shown in FIG. 15) is disposed on second side 1228 of buffer tank 1204 (similar to first conductor 204 in the embodiment shown in FIG. 2). A second conductor 1232 is disposed on a second side 1228 of buffer tank 1204

(also similar to second conductor 208 in the embodiment shown in FIG. 2).

A first electrode 1236, shown in FIG. 12 by a dashed line, is disposed within electrophoresis area 1212. First electrode 1236 extends downwardly into buffer solution 1242. First conductor 1504 provides a conductive path between a first terminal of a battery (not shown) housed in a recess 1246 and first electrode 1236, as shown in FIG. 14. In the embodiment shown, a conventional nine-volt transistor battery may be used as the power source to provide the necessary electricity for performing electrophoresis. A second electrode 1304 and a portion of second conductor 1232 is disposed on the bottom side of lid 1208. When lid 1208 is placed on top of buffer tank 1204, second conductor 1232 provides a conductive path from a second terminal ofthe battery to second electrode 1304, as shown in FIGs. 12-14.

For vertical electrophoresis, buffer solution must be disposed on either side ofthe gel to carry the electrical charge from an electrode disposed at the top ofthe gel to an electrode disposed at the bottom of each gel 1220. Thus, a tray 1250 is disposed in the top of buffer tank 1204. Tray 1250 has a Hp 1254 on either side to support tray 1250 in the top of buffer tank 1204. Tray 1250 has slots (not shown) formed therein for receiving the tops of gels 1220 therethrough. Buffer solution is added to tray 1250 so that it covers the tops of gels 1220.

Second electrode 1304 extends downwardly from the bottom of lid 1208 so that when lid 1208 is placed on top of buffer tank 1204, second electrode 1304 contacts the buffer solution in tray 1250 to complete the connection.

Thus, the buffer solution in tray 1250 has the opposite charge from buffer solution 1242 in the bottom of buffer tank 1204. The voltage difference between the buffer solutions causes charged protein or nucleic acid particles to travel down gels 1220 and separate. Because substantially vertical gel electrophoresis apparatus 1200 is made from an inexpensive material and operates using a conventional battery, apparatus 1200 can be packaged and sold in a cost-effective manner, similar to gel electrophoresis apparatus 100, 1100 and 2200.

While the invention has been particularly shown and described with reference to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope ofthe invention.

Claims

What Is Claimed Is:

1. An electrophoresis apparatus, comprising: a buffer tank defining, on a first side, an electrophoresis area; a first electrode mounted on a first portion ofthe apparatus; a second electrode mounted on a second portion ofthe apparatus; a lid configured to mate with said buffer tank to enclose said electrophoresis area; and a battery for supplying electricity to said first and second electrodes to cause a voltage differential between said first and second electrodes.

2. The apparatus of claim I, wherein said battery is disposed on the apparatus.

3. The apparatus of claim 1 , wherein said battery is disposed in a recess formed in said buffer tank.

4. The apparatus of claim 1, wherein said battery is disposed in a recess formed in said lid.

5. The apparatus of claim 1 , wherein a portion of a conductor is disposed on said lid so that when said lid is placed on top of said buffer tank said conductor completes an electrical connection between said battery and said first electrode.

6. The apparatus of claim 1 , further comprising a gel disposed in said buffer tank ofthe apparatus.

7. The apparatus of claim 1 , wherein said gel comprises agarose.

8. The apparatus of claim 1, wherein said gel comprises polyacrylamide.

9. The apparatus of claim 1 , further comprising a plurality of gels disposed in said buffer tank ofthe apparatus.

10. The apparatus of claim 1 , further comprising a buffer solution disposed in said buffer tank ofthe apparatus.

11. The apparatus of claim 1 , further comprising an open-celled, foam pad capable of being saturated with buffer solution and disposed in said buffer tank, said open-celled, foam pad disposed adjacent at least one of said first and second electrodes.

12. The apparatus of claim 1 , further comprising: a gel disposed in said buffer tank ofthe apparatus; an open-celled, foam pad capable of being saturated with buffer solution and disposed in said buffer tank, said open-celled, foam pad disposed adjacent at least one of said first and second electrodes; and a buffer solution disposed in said buffer tank ofthe apparatus.

13. The apparatus of claim 1 , further comprising: a first conductor to provide a first conductive path from said battery to said first electrode; and a second conductor to provide a second conductive path from said battery to said second electrode.

14. The apparatus of claim 1, wherein the electrophoresis apparatus is a substantially horizontal gel electrophoresis apparatus.

15. The apparatus of claim 1 , wherein the electrophoresis apparatus is a substantially vertical gel electrophoresis apparatus.

16. The apparatus of claim 1 , wherein the electrophoresis apparatus is an electroblotter apparatus.

17. The apparatus of claim 16, further comprising: a transfer package disposed in said buffer tank.

18. The apparatus of claim 17, wherein said transfer package comprises: a first blotter pad and a second blotter pad; a first filter disposed adjacent said first blotter pad and a second filter disposed adjacent said second blotter pad; a transfer membrane disposed between said first filter and said second filter; and a gel disposed adjacent said transfer membrane and between said first filter and said second filter.

19. The apparatus of claim 12, wherein said buffer tank comprises a registration key formed therein for orienting wells formed in said gel with respect to said buffer tank.

20. The apparatus of claim 1, wherein said lid is heat sealed to said buffer tank for shipping and storage ofthe apparatus.

21. The apparatus of claim 1, wherein said first and second electrodes are made from metal tape, said tape having an adhesive on one side to allow said first and second electrodes to be securely attached to the apparatus.

22. The apparatus of claim 1, wherein said lid is made from a transparent material.

23. The apparatus of claim 1, wherein said first and second electrodes are made from nickel.

24. The apparatus of claim 1 , wherein said first and second electrodes are made from stainless steel.

25. The apparatus of claim 1, wherein said first and second electrodes are deposited onto the apparatus by vapor deposition.

26. The apparatus of claim 1 , wherein said buffer tank is thermoformed from at least one of the following plastic materials: polyvinyl chloride, pheny lethyl thiogalactoside, and polystyrene.

27. The apparatus of claim 1, wherein said first and second electrodes are disposed on the apparatus in a manner permitting removal of said first and second electrodes for replacement.

28. The apparatus of claim 1 , wherein said buffer tank is molded from a sheet of plastic material to form a unitary buffer tank.

29. An electrophoresis apparatus, comprising: a buffer tank defining, on a first side, an electrophoresis area; a first electrode mounted on a first portion ofthe apparatus; a second electrode mounted on a second portion ofthe apparatus; and an open-celled, foam pad capable of being saturated with buffer solution and disposed in said buffer tank, said open-celled, foam pad disposed adjacent at least one of said first and second electrodes.

30. The apparatus of claim 29, further comprising: a power source for supplying electricity to said first and second electrodes to cause a voltage differential between said first and second electrodes.

31. The apparatus of claim 29, further comprising: a lid configured to mate with said buffer tank to enclose said electrophoresis area.

32. The apparatus of claim 30, wherein said power source is a battery.

33. The apparatus of claim 29, wherein the electrophoresis apparatus is a substantially horizontal gel electrophoresis apparatus.

34. The apparatus of claim 29, wherein the electrophoresis apparatus is a substantially vertical gel electrophoresis apparatus.

35. The apparatus of claim 29, wherein the electrophoresis apparatus is an electroblotter apparatus.

36. An electrophoresis apparatus, comprising: a buffer tank defining, on a first side, an electrophoresis area configured to contain a cast gel and buffer solution and defining, on a second side, a recess for receiving a battery; a first electrode mounted on a first portion ofthe apparatus; a second electrode mounted on a second portion ofthe apparatus; a first conductor for providing a first conductive path from said recess to said first electrode; and a second conductor for providing a second conductive path from said recess to said second electrode.

37. The apparatus of claim 36, further comprising: a lid configured to mate with said buffer tank to enclose said electrophoresis area.

38. The apparatus of claim 36, further comprising: an open-celled, foam pad capable of being saturated with a buffer solution and disposed in said buffer tank.

39. The apparatus of claim 36, wherein the electrophoresis apparatus is a substantially horizontal gel electrophoresis apparatus.

40. The apparatus of claim 36, wherein the electrophoresis apparatus is a substantially vertical gel electrophoresis apparatus.

41. The apparatus of claim 36, wherein the electrophoresis apparatus is an electroblotter apparatus.

42. An electrophoresis apparatus, comprising: a buffer tank defining, on a first side, an electrophoresis area; a first electrode mounted on said first portion ofthe apparatus; a lid, configured to mate with said buffer tank and to enclose said electrophoresis area; and a second electrode mounted on a first side of said lid so that when said lid mates with said buffer tank said second electrode is opposite said first electrode, wherein said lid and said buffer tank each have a tab formed in a periphery so that when said lid and said buffer tank are matingly combined, said tabs are opposite one another, said tab of said buffer tank configured to mateably engage a first tab receiving area of an adaptor, and said tab of said lid configured to mateably engage a second tab receiving area of an adaptor to provide a conductive path from said first and second electrodes to a power source.

43. A disposable electrophoresis kit, comprising: a. a unitary buffer tank molded from a sheet of plastic, said buffer tank defining, on a first side, an electrophoresis area; b. a first electrode mounted on a first portion ofthe kit, c. a second electrode mounted on a second portion of the kit; d. a first conductor for providing a first conductive path from a power source to said first electrode; e. a second conductor for providing a second conductive path from said power source to said second electrode; f. a lid, configured to mateably engage said buffer tank; g. a pre-cast gel; and h. at least one open-celled, foam pad capable of being saturated with a buffer solution disposed in said buffer tank adjacent to at least one of said first and second electrodes.

44. The disposable electrophoresis kit of claim 43 , wherein said power source is a battery.

45. A method for separating a protein or nucleic acid sample in an electrophoresis apparatus, said method comprising the steps of: (a) obtaining the apparatus of claim 1 ;

(b) applying a sample to a gel; and

(c) creating a voltage differential to allow separation of said sample in said gel.

46. A method for transferring a protein or nucleic acid sample from a gel to a membrane, said method comprising the steps of:

(a) obtaining the apparatus of claim 1;

(b) disposing in said buffer tank in close proximity a gel containing said protein or nucleic acid with a transfer membrane;

(c) creating a voltage differential to allow transfer of the protein or nucleic acid from the gel to the transfer membrane.