A GEL CASTING ASSEMBLY FOR HORIZONTAL GEL ELECTROPHORESIS FIELD OF THE INVENTION:

This invention relates to a novel gel casting assembly for gel electrophoresis. More particularly it relates to the novel design of the gel casting assembly which enable sequential electrophoresis in two different orientations. BACKGROUND OF THE INVENTION:

Gel electrophoresis is the method of separating biomolecules within a gel matrix on the basis of their molecular weight and charge under the influence of electric current. The separation is carried out by running a mixture of samples through a gel, wherein the pores in the mesh or matrix allows the movement of the molecules based on their molecular size. The higher molecular weight molecules is resisted in their movement in comparison to the lower molecular weight with pore size being a decisive factor. Higher the concentration of the gel material, the smaller is the pore size and thus smaller biomolecules are better separated than the medium and larger biomolecules. The gel electrophoresis is broadly classified into horizontal and vertical electrophoresis.

Many suppliers specializes in providing the entire assembly for carrying out electrophoresis. A particular horizontal gel electrophoresis assembly will comprise of an electrophoretic tank unit with a lid, a casting gel tray, multichannel combs, connecting plug for positive & negative electrode and a power pack.

Till date, the units for casting gels are provided which have their own set of problems. These units are El shaped devices which are confined for only one orientation electrophoretic run. The length and width of gel is predetermined enabling only single directional gel electrophoresis. The number of samples that could be analysed are dependent on width of devices whereas the length determine the resolution of analytes from sample. The other open ends of the device need to be sealed for gel casting and liquid leakage prevention till complete gelation. Said open ends are sealed using cello tape. In this type of gel casting device and in all the possible options available today, the comb is always aligned parallel to the width of the gel device. If more numbers of samples are to be analysed, either multiple analyses need to be carried out resulting in multiple gel to be compared for deriving conclusions or another gel casting device with extended width need to be bought and used. Again when the samples need to be run for a very a short distance for resolution, still the regular device has to be used resulting in wasting of gel or another gel casting device with short length need to be bought and used. Ultimately the researcher, analyst need to buy multiple gel casting devices in various lengths and widths. Thus, the available choices were tedious and not at all cost effective. The current invention provides a gel casting assembly where the comb can be aligned parallel to the width as well as the length of the gel casting unit. It provides various advantages in cost effective manner. Hereinafter, the assembly and its utility is explained in detail.

SUMMARY OF THE INVENTION:

In one aspect the invention provide a gel casting assembly for gel electrophoresis which comprises

a) a rectangular planar body with an upper surface; a basal surface; four side surfaces connecting said upper and basal surfaces and means for attaching detachable side walls,

b) four side walls having means for resting one or more comb and capable to detachably attach with said rectangular planar body to remain perpendicular to either upper or basal surface of said planar body through said means for attaching to create a rectangular mould for casting gel, wherein after attachment of said side walls to said rectangular planar body, each side wall forms a pair with its parallel side wall and allow resting of one or more comb on them in a direction parallel to the other side walls when said other sidewalls do not have any comb resting on them.

In another aspect it discloses a process of carrying out two gel electrophoresis sequentially in two different orientations using a single gel casting assembly. The process comprises following steps:

a) taking a gel casting assembly containing

a rectangular planar body with an upper surface; a basal surface; four side surfaces connecting said upper and basal surfaces and means for attaching detachable side walls,

four side walls having means for resting one or more comb and capable to detachably attach with said rectangular planar body to remain perpendicular to either upper or basal surface of said planar body through said means for attaching to create a rectangular mould for casting gel;

b) attaching said four side walls to said rectangular planar body;

c) resting one or more comb on first pair of parallel side walls in a direction parallel to the other two side walls;

d) pouring a molten gel in said rectangular mould and allowing it to solidify;

e) removing the one or more comb and said other two side walls and putting the assembly in electrophoretic tank having buffer; f) loading the sample in the wells created by removal of the one or more comb and carrying out electrophoresis by supplying electric charge at the sides of removed side walls through anode and cathode; g) removing the gel casting assembly from the electrophoretic tank after completion of electrophoresis; h) cleaning the gel casting assembly to repeat steps a) to g) but with using second pair of parallel side walls in step c) for resting one or more comb and accordingly removing the other side walls in step e). BRIEF DISCRETION OF ACCOMPANYING DRAWINGS:

All aspects and advantages of the present invention will become apparent with the description of the preferred non limiting embodiment, when read together with the accompanying drawings, in which: FIG la and lb illustrate the exemplary embodiments for the conventional gel casting device in U shape, wherein the dotted line in FIG lb indicate the sealants covering the open ends of said device;

FIG 2 illustrate an exemplary embodiment for a rectangular planar body with an upper surface (200a), a basal surface (200b) and four side surfaces (20 G, 202’, 203’ and 204’);

FIG 3 illustrate an exemplary embodiment for the rectangular planar body (200) with groves (201, 202, 203 and 204) adjacent and parallel to four side surfaces of said planar body;

FIG 4 illustrate an exemplary embodiment for the gel casting assembly (300) having rectangular planar body (200) attached with four side walls (301,302, 303,304), each having groves for resting one or more comb (305 to 314);

FIG 5 illustrate an exemplary embodiment for the gel (401) casted in gel casting assembly and the comb (402) mounted parallel to the width of gel casting assembly to allow the length wise run of samples; FIG 6 illustrate an exemplary embodiment for the gel (501) casted for length wise run of samples, with the wells (502) for sample loading formed due to the comb (402); FIG 7 illustrate an exemplary embodiment for the gel (601) casted in gel casting assembly and the comb (602) mounted parallel to the length of gel casting assembly to allow the width wise run of samples;

FIG 8 illustrate an exemplary embodiment for the gel (701) casted for width wise run of samples, with the wells (702) for sample loading formed due to the comb (502);

DETAILED DESCRIPTION OF THE INVENTION:

As mentioned, there remains a long felt need to decipher a device or assembly, which is unequivocally efficient and effective in providing ease and cost effective solution for conducting horizontal electrophoresis. The present invention tries to address this long felt need.

The current invention provides a gel casting assembly where the comb can be aligned parallel to the width as well as the length of the gel casting unit. It enables to carry out gel electrophoresis in any one orientation at a time but with choice of either in length-wise or width-wise orientation. It also enable the electrophoresis to be carried out sequentially in two different orientations using same gel casting assembly. It provides multiple sample analysis for short run samples by aligning comb parallel to the length and also allow long run of samples by aligning comb parallel to the width. In fact it also enable the researcher to study separation of the components at different lengths using two different orientations of same gel casting unit by keeping all other electrophoretic parameters same. According to the first aspect of the invention, this assembly mainly comprises a) a rectangular planar body with an upper surface; a basal surface; four side surfaces connecting said upper and basal surfaces and means for attaching detachable side walls, and

b) four side walls having means for resting one or more comb and capable to detachably attach with said rectangular planar body to remain perpendicular to either upper or basal surface of said planar body through said means for attaching to create a rectangular mould for casting gel.

After attachment of said side walls to said rectangular planar body, each side wall forms a pair with its parallel side wall and allow resting of one or more comb on them in a direction parallel to the other side walls when said other sidewalls do not have any comb resting on them.

FIG 2 illustrate an exemplary embodiment for a rectangular planar body with an upper surface (200a), a basal surface (200b) and four side surfaces (20 G, 202’, 203’ and 204’);

The four detachable side walls are preferably planar and rectangular in shape and each has means for resting one or more comb.

The rectangular planar body and four side walls are preferably made from a chemically inert, UV transparent, scratch resistant and autoclavable material. Preferably such material is selected from UV transparent acrylic material, OP-l grade cast acrylic material, silicone material, and any other material being chemically inert, UV transparent, scratch resistant and autoclavable.

This assembly enables the casting of gel in rectangular dimension and give choice of sample run in length wise or width wise orientation by aligning the comb in specific direction while gel casting and then removing the comb and unwanted side walls after gelation is complete. Due to availability of more than one comb resting means, it is possible to rest multiple combs for analysing multiple short run samples in a single experiment.

Various means can be adopted for attaching the detachable side walls to the rectangular planar body. Examples of such means are adhesive, magnets and hinges. Alternatively, the detachable side walls may be attached to the rectangular planar body with holes and detachable threaded or unthreaded pins. The pins could be permanently affixed to or protruding from any of the component while another component being having holes to accommodate it inside and keep the sidewalls temporarily attached to the rectangular planar body. The removable pins could also be used by providing holes in both components.

In one embodiment, the means for attaching detachable side walls to rectangular planar body are protrusion and socket fixture arrangement made between side walls and side surfaces of said rectangular planar body.

In yet another embodiment, the groves are made on upper or basal surface of rectangular planar body. These grooves accommodate the side walls basal portion in them. Preferably these grooves are made adjacent and parallel to four side surfaces of said planar body. Alternatively groves can be made on side walls to accommodate the side surfaces of rectangular planar body in them.

In any types of arrangement for attaching detachable side walls to the rectangular planar body, the precaution is taken that the four side walls create a leakage proof rectangular mould for casting gel over the rectangular planar body.

The means for resting one or more comb on the side walls are selected from groves, magnets and hooks. The grooves being the simplest means is preferred. Nevertheless, the magnets and hooks can also be implemented for this purpose without deviating from the spirit of this invention. FIG 3 further illustrate an exemplary embodiment where groves (201, 202, 203 and 204) are created on upper or basal surface of rectangular planar body (200).

FIG 4 further illustrate an exemplary embodiment for a gel casting assembly (300) having rectangular planar body (200) attached with four side walls (301, 302, 303 and 304), each having groves for resting one or more comb. The side walls 301 and 302 forms a pair with each other and allow resting of one or more comb on them in a direction parallel to side walls 303 and 304 using grooves made therein for width wise sample run. Similarly the side walls 303 and 304 forms a pair with each other and allow resting of one or more comb on them in a direction parallel to side walls 301 and 302 using grooves made therein for length wise sample run.

FIG 5 further illustrate an exemplary embodiment where the gel (401) is casted in gel casting assembly and the comb (402) is mounted parallel to the width of gel casting assembly to allow length wise run of samples.

FIG 6 further illustrate an exemplary embodiment where the gel (501) is casted for length wise run of samples, having the wells (502) for sample loading formed due to the comb (402).

FIG 7 further illustrate an exemplary embodiment where the gel (601) is casted in gel casting assembly and the comb (602) is mounted parallel to the length of gel casting assembly to allow width wise run of samples. FIG 8 further illustrate an exemplary embodiment where the gel (701) is casted for width wise run of samples, having the wells (702) for sample loading formed due to the comb (602).

On the basis of the number of samples and orientation for the run, the researcher/user can place the combs in the respective orientation. If the combs are aligned parallel to the width of gel casting assembly [FIG 5] then the electrophoretic run is in the length - wise orientation and maximum distance for sample run is provided [FIG 6] whereas combs when aligned parallel to the length of gel casting assembly [FIG 7], it result in width wise orientation [FIG 8] and provide for minimal distance for sample run in gel.

Here the maximum distance of sample run hold true when only a single comb is mounted. However, another advantage of width-wise gel orientation is that more number of wells can be created or accommodated than length-wise orientation thus more samples can be run simultaneously.

In second aspect the invention provides a process of carrying out two gel electrophoresis sequentially in two different orientations using a single gel casting assembly. This process comprises

a) taking a gel casting assembly containing

a rectangular planar body with an upper surface; a basal surface; four side surfaces connecting said upper and basal surfaces and means for attaching detachable side walls,

four side walls having means for resting one or more comb and capable to detachably attach with said rectangular planar body to remain perpendicular to either upper or basal surface of said planar body through said means for attaching to create a rectangular mould for casting gel;

b) attaching said four side walls to said rectangular planar body;

c) resting one or more comb on first pair of parallel side walls in a direction parallel to the other two side walls;

d) pouring a molten gel in said rectangular mould and allowing it to solidify;

e) removing the one or more comb and said other two side walls and putting the assembly in electrophoretic tank having buffer; f) loading the sample in the wells created by removal of the one or more comb and carrying out electrophoresis by supplying electric charge at the sides of removed side walls through anode and cathode; g) removing the gel casting assembly from the electrophoretic tank after completion of electrophoresis;

h) cleaning the gel casting assembly to repeat steps a) to g) but with using second pair of parallel side walls in step c) for resting one or more comb and accordingly removing the other side walls in step e). The gel, buffer and samples in both electrophoresis can be same or different.

It provide an added advantage when the researcher or analyst need to study separation of the components at different lengths in gel, he/she can keep all the parameters same and change only the sample run orientations using same gel casting assembly as explained above.

Same electrophoretic buffer can be used for different gels and different samples too. Similarly by using same gel casting assembly, it is possible to analyse same samples in different gels in different orientations keeping same or different electrophoretic buffer.

In fact the gel, buffer and samples in both electrophoresis may be of same concentration or different concentration. Examples:

Preparing a gel mould and casting gel into it for maximum distance sample run

For casting an agarose gel with wells using the current casting gel assembly, firstly the gel slab (200) is taken [FIG 3] and four walls (301, 302, 303 & 304) are slided over the groves (301, 302, 303 & 304). The walls are locked into each other resulting in rectangular mould for casting gel [FIG 4] The walls on all the corners provide for a confinement for casting and preventing gel leakage.

For casting a gel to allow maximum distance sample run, here being the entire length of the casting assembly, combs are mounted on to groves (305 and 308) [FIG 4 & 5] The heated agarose gel solution of a particular concentration (401) prepared in electrophoretic buffer is poured into the intact gel casting assembly mounted with a comb (402). The heated agarose solution is allowed to cool at room temperature resulting in gelation and formation of an agarose matrix (501). The comb is carefully removed resulting in the well formation (502) for sample loading [FIG 6] The walls (301 & 302) are removed by sliding out through the grooves. The gel casting assembly with the agarose gel is placed in an electrophoretic tank with electrophoretic or run buffer & samples are loaded. After closing the lid and connecting the power plug, selection of the orientation of current is decided with the provision in the power pack, the samples are allowed to run.

Preparing a gel mould and casting gel into it for minimum distance sample run For casting an agarose gel with wells using the current casting gel assembly, the similar procedure as mentioned in above example was followed to create a rectangular mould for casting gel [FIG 4] using same gel casting assembly.

For casting a gel with minimal distance sample run, here being the entire width of the casting assembly, combs are mounted on to groves (311 & 314) [FIG 4 & 7] The heated agarose gel solution (601) of a particular concentration prepared in electrophoretic buffer is poured into the intact gel casting assembly mounted with the combs (602). The heated agarose solution is allowed to cool at room temperature resulting in gelation and formation of an agarose matrix (701). The combs are carefully removed resulting in the well formation (702) for sample loading [FIG 8] The walls (303 & 304) are removed by sliding out through the grooves. The gel casting assembly with the agarose gel is placed in an electrophoretic tank with electrophoretic or run buffer and samples are loaded. After closing the lid and connecting the power plug, selection of the orientation of current is decided with the provision in the power pack and anodes and cathodes, the sample are allowed to run.

In all the cases, the user has the choice of adding more than one comb depending on the number of samples that needs to be run at a given time.

The foregoing description of the specific embodiments will so fully reveal the general nature of the embodiments herein that others can, by applying current knowledge, readily modify and/or adapt for various applications such specific embodiments without departing from the generic concept, and, therefore, such adaptations and modifications should and are intended to be comprehended within the meaning and range of equivalents of the disclosed embodiments. It is to be understood that the phraseology or terminology employed herein is for the purpose of description and not of limitation. Therefore, while the embodiments herein have been described in terms of preferred embodiments, those skilled in the art will recognize that the embodiments herein can be practiced with modification within the spirit and scope.