JPS60164242A - Polyacrylamide gel film for electrophoresis - Google Patents

Abstract

PURPOSE:To simplify separating operation by polymerizing nonwoven fabric under the state it is immersed in the aqueous solution of the mixture of acrylamide monomer and NN'-methylene bisacrylamide, and forming polyacrylamide gel so as to surround the unwoven fabric. CONSTITUTION:The unwoven fabric, which is formed by polyester and the like, is immersed in the aqueous solution of the mixture of acrylamide monomer and NN'-methylene bisacrylamide. Under this state, the acrylamide monomer and the NN'-methylene bisacrylamide are polymerized, and polyacrylamide gel is formed so as to surround the unwoven fabric. The unwoven fabric is washed and dried, and polyacrylamide gel film for electrophoresis is formed. The film has the following characteristics: economic burdens are few; it can be prepared beforehand; separating operation can be simplified; it does not include salts, which impede the reproducibility of separation; its strength is large; and the thickness can be made thin.

Description

DETAILED DESCRIPTION OF THE INVENTION [Technical Field of the Invention] The present invention relates to a polyacrylamide gel film for electrophoresis.

[Technical background of the invention and its problems] Zone electrophoresis is an extremely effective and widely used method for separating proteins, enzymes, etc. Among them, the method using polyacrylamide gel requires the use of electricity during electrophoresis. As a result of almost no penetration phenomenon, it exhibits high separation power,
This is a highly useful method.

In electrophoretic separation operations using polyacrylamide gel, as seen in the disk method or slab gel method, the first step is to prepare an aqueous acrylamide/septumer solution in consideration of the separation conditions. 11 polymerizing it in a separation vessel to form a separation zone;
The second step is to place the sample material on the created separation zone and perform electrophoretic separation, and the third step is to stain the separation zone and confirm the separation state. is common.

Looking at the points that should be improved regarding this method, the first point is that it is necessary to prepare the gel prior to the separation operation. This is one of the operational complications and may hinder reduction of the time required for analysis. This problem can be solved by preparing a large number of separation containers and preparing and storing the gel in advance, but this involves a large economic burden.

Second, the gel contains salts added as polymerization promoters and polymerization catalysts in addition to the buffer added to prepare the environment. The presence of these extra salts must not only be taken into consideration during normal electrophoresis, but also in cases such as isocentral electrophoresis, their presence may have a negative effect on separation. Therefore, there are cases where the separation operation becomes complicated, such as the need to perform pre-phoresis and remove them prior to the separation operation 1'1°.

The third factor is the thickness and strength of the gel.

After electrophoresis is completed, the gel is removed from the container and stained to confirm the location of the sample and other conditions. Therefore, the gel needs to be strong enough to withstand this operation, and for this reason, it needs to be thick to a certain extent, which results in an increase in the time required for the staining operation.

The fourth is dimensional stability. The size of the gel [well, it varies greatly depending on the water content of the gel, so the staining line f
[The size of the gel after staining may vary depending on the
There are also cases where it is difficult to accurately compare migration positions between gels.

[Object of the Invention] The present invention is based on the above-mentioned pulling foil, and can be prepared in advance with less economical burden, simplifying the separation operation, and improving the reproducibility of the separation. The purpose of the present invention is to obtain a polyacrylamide gel film for electrophoresis that does not contain salts that can interfere with electrophoresis, has high strength, is small in thickness, and has good dimensional stability.

[Summary of the Invention] The polyacrylamide gel film for electrophoresis of the present invention is produced by immersing a nonwoven fabric made of polyester or the like in an aqueous solution of a mixture of an acrylamide monomer and NN'-methylenebisacrylamide. and NN'-methylenebisacrylamide to form a polyacrylamide gel surrounding a nonwoven fabric, which is then washed and dried.

[Embodiments of the Invention] The polyacrylnoamide gel film for electrophoresis of the present invention was prepared by first preparing 4.85 g of acrylamide 7-mer;
N,N'-methylenebisacrylamide 0.15 (1
, N, N, N', N'-tetramethylethylenediamine 10111 g, ammonium persulfate ioomg,
1C) Make an aqueous solution with a proportion of OmN dissolved in water,
Degas for 1 j under reduced pressure. Similarly, at 11, r, a 0.5 mm thick rubber band is used as a spacer between two glass plates, with a distance of lυ, and an extremely thin non-woven polyester fabric is inserted into the gap. Have an IC ready. The previously prepared aqueous solution was poured into this gap by 21 people, being careful not to leave any air, and the glass plate was wrapped with polyvinylidene chloride film, etc. to block air and left to stand.
Allow polymerization to complete. After polymerization is complete, push a spatula-like object between the glass plates, remove the glass plate, and remove the gel.

After thoroughly washing the gel with pure water to remove the salts contained therein, spread it out on a board made of a material such as polyethylene or Teflon that will naturally peel off once the gel dries. , a film-like polyacrylamide gel film is obtained by drying.

This gel film has high tensile strength, can be handled like paper, and can be stored for long periods under normal environmental conditions. When this gel film is immersed in water or a buffer solution for about 30 minutes, it becomes a gel with a high mechanical strength and approximately the same area as when dry. This gel can be used as an electrophoresis zone, as can buffer-impregnated filter paper or hillose acetate film.

This gel naturally does not contain polymerization accelerators or other salts that may have a negative effect on separation, so it does not require pre-phoresis or other preliminary operations during isoelectric focusing, and has excellent separation performance during electrophoresis. is similar to normal polyacrylamide gel without drying. By using this gel, a gel with the necessary conditions can be used at any time, and the operation of electrophoresis using polyacrylamide gel is comparable to that of filter paper electrophoresis.
Simplified.

This gel has extremely high tensile strength and there is almost no risk of breakage, so it is easy to handle and can be made as thin as a non-woven fabric. It is appropriate that the Q is 3mm to 0.5In lit. In this case, the time required for staining and other treatments can be reduced to a few minutes compared to the case of a 1 mm thick gel.
At the same time, the high dimensional stability made it easy to compare electrophoretic images after staining.

Further, this gel film can be cut with scissors or the like, and can be printed using a common writing instrument such as a pencil. Therefore, prior to the scheduled electrophoresis procedure 1, cut the dry gel film to the required size -C to avoid waste;
In addition, after writing the type of sample, addition position, direction of current, etc. on the gel film, it is immersed in a buffer solution to restore the gel state, and electrophoresis is performed according to the written conditions to analyze the electrophoretic image. This is extremely effective as it can avoid confusion during the process.

The present invention is not limited to the above embodiments. In other words, the materials used as the core of the polyacrylamide gel film include polyester nonwoven fabric, nylon, polypropylene, etc., which maintain high tensile strength even when wet, are chemically stable, and have good dimensional stability. Non-woven fabrics and cloth using fibrous materials, or porous films with these properties can be used, and gel formulations include acrylamide monomer, N,N'-methylenebis, etc. Acrylamide, N, N, N', N'-tel
The ratio of methylethylenediamine to ammonium persulfate can be changed as desired, and the same effect can be obtained by using a polymerization initiator such as riboflavin instead of ammonium persulfate.

The container used for polymerization of the gel is not limited to a glass plate, but may also be a plastic plate or a box made of glass, plastic, etc. Regarding the method of drying the gel, In addition to drying the gel by placing it on a plate that can be easily peeled off after drying, fixing the surrounding area to a frame to prevent the gel from wrinkling is also a method of drying, and regarding drying conditions, in addition to natural drying, vacuum Good results can also be obtained by methods such as drying and heating drying.

"Effects of the Invention" As is clear from the above, the gel film of the present invention can be manufactured in advance and stored under normal environmental conditions, thereby simplifying the separation operation. Furthermore, since it does not contain salts etc., separation with good reproducibility is possible.

Application agent Patent attorney Kikuike Department 5

Claims (1)

[Claims] A nonwoven fabric made of polyester or the like is immersed in an aqueous solution of a mixture of an acrylic 7-mide monomer and NN'-methylenebisacrylamide, and the acrylamide monomer and NN'-methylenebisacrylamide are polymerized to surround the nonwoven fabric. A polyacrylamide gel film for electrophoresis, characterized in that it is formed by forming a polyacrylamide gel, washing and drying the nonwoven fabric.