JP2002088652A - Method for introducing hydrophilic functional group into inner wall portion of hollow fiber and method for filling gel into hollow fiber - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for treating the inner wall of a hollow fiber so that a gel to be filled into the hollow portion is physically or chemically fixed to the inner wall of the hollow fiber, and to provide a method for filling the gel into the hollow portion of the hollow fiber, which is suitable for use in micro-arrays for capillary electrophoresis and DNA analysis. SOLUTION: This method for introducing the hydrophilic functional groups into the inner wall portion of the hollow fiber comprises adhering the impermeable solution of a polymer having the hydrophilic groups to the inner wall portion of the hollow fiber. The method for filling the gel into the hollow portion of the hollow fiber is characterized by adhering the polymer to the inner wall portion and then filling the gel into the hollow portion.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for introducing a hydrophilic functional group into the inner wall portion of a hollow fiber and a method for stably filling a hollow portion of a hollow fiber with a gel. The gel-filled hollow fiber or capillary-like material prepared in the present invention is usefully used for microarrays for capillary electrophoresis and DNA analysis.

[0002]

2. Description of the Related Art As a method for filling a hollow fiber with a gel, Japanese Patent Application Laid-Open No.
No. 21694 has been proposed. However, the gel to be filled tends to peel off from the hollow fibers due to polymerization shrinkage that usually occurs during polymerization and fall off from the hollow fibers. Therefore, it has been difficult to use the hollow fiber filled with the gel for a microarray for capillary electrophoresis or DNA analysis.

[0003]

SUMMARY OF THE INVENTION An object of the present invention is to provide a method for treating the inner wall of a hollow fiber in which a gel filled in the hollow is physically or chemically fixed to the inner wall of the hollow fiber, and a capillary electrophoresis. And a method for filling a gel into a hollow portion of a hollow fiber suitable for use in a microarray for DNA analysis and DNA analysis.

[0004]

Means for Solving the Problems The present inventors have conducted intensive studies in order to solve the above-mentioned problems, and as a result, in a method for treating the inner wall of a hollow fiber, the hollow fiber has a hydrophilic group capable of penetrating into the inner wall. After depositing the polymer solution, the present inventors have found that the gel filled in the hollow portion can be provided on the inner wall portion of the hollow fiber by providing the polymer on the inner wall portion, thereby providing a place where the gel is physically fixed. .

That is, the present invention provides a method for treating a hollow fiber inner wall, wherein a polymer having a hydrophilic group is adhered to the inner wall, and a method for introducing a functional group into the hollow fiber inner wall. Filling a hollow portion of a hollow fiber in which a polymer having a functional group adheres to an inner wall portion of the hollow fiber with a gel-forming polymerizable monomer, and polymerizing the monomer to fill the hollow portion of the hollow fiber with a gel. The way.

[0006]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, embodiments of the present invention will be described. The hollow fiber used in the present invention may be a porous hollow fiber or a non-porous hollow fiber. Examples thereof include nylon 6, nylon 66, various polyamide-based fibers such as aromatic polyamide, polyethylene terephthalate, polybutylene terephthalate, and the like. Various polyester fibers such as polylactic acid, polyglycolic acid, and polycarbonate,
Various fibers of acrylic such as polyacrylonitrile, various fibers of polyolefin such as polyethylene and polypropylene, various fibers of polymethacrylate such as polymethyl methacrylate, various fibers of polyvinyl alcohol, various fibers of polyvinylidene chloride, poly Examples include vinyl chloride fibers, various polyurethane fibers, phenol fibers, fluorine fibers made of polyvinylidene fluoride, polytetrafluoroethylene, and the like, and various polyalkylene paraoxybenzoate fibers.

For capillary electrophoresis, a transparent material is preferable in order to irradiate detection light from outside the capillary, and a hollow fiber or a capillary (hereinafter referred to as a hollow fiber) made of a methacrylate resin represented by polymethyl methacrylate is used as a material. ) Are preferably used.

The outer diameter of the hollow fiber used is 2 mm or less, preferably 1 mm or less. Further, the inner diameter is preferably 0.02 mm or more. For capillary electrophoresis, a relatively thick hollow fiber is preferable because of easy handling. In the case of a porous hollow fiber, the hollow fiber inner wall portion that is the object of the present invention includes the inner wall surface and a portion that has entered the interior from the inner wall surface. In the case of non-porous hollow fibers, it refers to the inner wall surface.

Also, a DNA chip using a fiber array using a hybridization technique (Japanese Patent Application No. 11-84100).
No.) for the hollow fiber treated by the method of the present invention,
Gel-filled hollow fibers can also be made. In that case,
Immobilizing the probe DNA on the gel-filled hollow fiber of the present invention,
A number of fibers are arranged, hardened with resin, and sliced at right angles to the fiber axis to produce chips. In such applications, it is necessary that the number of fibers per unit area is large, and the outer diameter of the fibers is preferably thinner, preferably 0.5 mm or less, more preferably 0.1 mm or more and 0.3 mm or less. is there.

Further, in a DNA chip using a fiber array, a tension is applied to the fibers at the stage of arrangement in order to maintain the regularity of the arrangement in the manufacturing method, so that a material having a high elastic modulus, for example, aromatic polyamide or methyl methacrylate is used. Fibers made of a methacrylic resin such as the above are preferred.

Since the gel-filled hollow fiber of the present invention is used for electrophoresis and DNA analysis as described above, the gel to be filled in the hollow portion and / or the porous portion contains water and water. Mainly polyacrylamide gel with affinity. Therefore, as the polymer having a hydrophilic group for treating the inner wall portion of the hollow fiber, a carboxylic acid group,
A polymer having a hydroxyl group, an amino group, a sulfonic acid group, an anionic group, or the like, or a nonionic polymer is preferable.

The monomers forming the polymer include:
Acrylic acid, methacrylic acid, 2-hydroxyethyl acrylate, 2-hydroxyethyl methacrylate, 2-
Hydroxypropyl acrylate, vinyl alcohol,
Acrylamide, methacrylamide, sodium styrene p-sulfonate, ethylene glycol and the like can be mentioned.

These monomers can be used alone or in combination of two or more. Further, a copolymer with another polymerizable monomer having an unsaturated double bond can also be used. Such monomers include (meth) acrylates, styrenes,
(Meth) acrylamide-based monomers and the like.

Examples of (meth) acrylate monomers include methyl methacrylate, ethyl methacrylate, n-propyl methacrylate, isopropyl methacrylate, n-butyl methacrylate, isobutyl methacrylate, t-butyl methacrylate, and methacrylic acid. Methacrylates such as isoamyl, lauryl methacrylate, phenyl methacrylate, benzyl methacrylate, cyclohexyl methacrylate, glycidyl methacrylate, 2-ethylhexyl methacrylate, and methyl acrylate, ethyl acrylate, n-propyl acrylate, and acrylic acid Isopropyl, n-butyl acrylate, isobutyl acrylate, t-butyl acrylate, isoamyl acrylate, lauryl acrylate, phenyl acrylate, benzyl acrylate, acrylic acid Kurohekishiru, glycidyl acrylate, acrylic acid esters such as 2-ethylhexyl acrylate.

Examples of the styrene monomer include:
o-methylstyrene, m-methylstyrene, p-methylstyrene, α-methylstyrene, 2,4,6-trimethylstyrene, p-methoxystyrene, α-methoxystyrene, p-1- (2-hydroxybutyl) styrene , P-
1- (2-hydroxypropyl) styrene, p-2-
(2-hydroxypropyl) styrene and the like.

Examples of acrylamide monomers include:
N-methyl (meth) acrylamide, N, N-dimethyl (meth) acrylamide, N-ethyl-N-methyl (meth) acrylamide, N, N-diethyl (meth) acrylamide, Nn-propyl (meth) acrylamide,
N-isopropyl (meth) acrylamide, Nt-butyl (meth) acrylamide, Ns-butyl (meth)
Acrylamide, Nn-butyl (meth) acrylamide, N-methyl-N-isopropyl (meth) acrylamide, N-methyl-Nn-propyl (meth) acrylamide, N-ethyl-N-isopropyl (meth) acrylamide, N-ethyl-NN-propyl (meth) acrylamide, N, N-di-n-propyl (meth) acrylamide and the like are exemplified.

Using these monomers, a polymer having a hydrophilic group for treating the inner wall of the hollow fiber can be obtained by ordinary solution polymerization or suspension polymerization. As a solvent used for treating the inner wall of the hollow fiber, any solvent can be used as long as it dissolves the polymer and can penetrate into the inner wall of the hollow fiber. Preferred solvents include alcohols such as methanol, ethanol and propanol, acetone, toluene, ethyl acetate and the like. The degree of treatment of the inner wall can vary depending on the polymer concentration in the polymer solution and the like. The polymer concentration is preferably in the range of 50% or less, and more preferably in the range of 1 to 30%.

Next, a specific processing method will be described.
The distal end of the extended portion of the hollow fiber is immersed in a solution containing a polymer having a hydrophilic group, inhaled, and the polymer solution is introduced into the inner wall of the hollow fiber to attach the polymer. The polymer solution may be introduced by any method. For example, a method in which the open end of a hollow fiber whose one end is closed is immersed in the polymer solution and the hollow fiber is filled with the solution by suction can be exemplified. When the polymer is adhered by the above method and left, the hollow portion is clogged. Therefore,
It is preferable that the polymer solution is partially released again after the suction once.

Next, a method for filling the hollow portion of the hollow fiber obtained by the above treatment with gel will be described. Examples of the gel-forming polymerizable monomer solution to be filled include monomers that form the polymer used in the treatment of the inner wall portion of the hollow fiber. As the polymerization initiator, an azo-based, peroxide-based, or redox-based initiator that can be dissolved in a solvent to be used can be used. Examples include 2,2'-azobisisobutyronitrile, 2,2'-azobis (2-methylbutyronitrile) isobutyronitrile, benzoyl peroxide, and benzoyl peroxide-dimethylaniline. . Examples of the cross-linking agent necessary for gel formation include acrylamide monomers having two or more functionalities, such as N, N'-methylenebisacrylamide,
N '-(1,2-dihydroxyethylene) -bisacrylamide, N, N'-diallyl tartardiamide, N,
N'-cystamine-bisacrylamide, N-acryloytris (hydroxymethyl) aminomethane and the like are preferred.

Particularly, an acrylamide gel is preferable for DNA analysis. The acrylamide monomer concentration is preferably in the range of 2 to 20%, and polymerization is performed by adding a crosslinking agent and a polymerization initiator to an aqueous solution. The method of filling the hollow portion of the hollow fiber with the monomer solution is generally a vacuum suction method, but is not limited thereto.

[0021]

EXAMPLES Example 1 Synthesis of Methyl Methacrylate-Methacrylic Acid Copolymer A toluene solution A having the following composition was prepared, and a methyl methacrylate-methacrylic acid copolymer used in the treatment was synthesized. . The toluene solution A was polymerized in a reaction vessel at 70 ° C. for 6 hours under a nitrogen atmosphere, and then a solution diluted three times with methyl ethyl ketone was poured into seven times the amount of hexane to obtain a methyl methacrylate-methacrylic acid copolymer. Combined (methyl methacrylate / methacrylic acid = 86 mol)
% / 14 mol%).

<Toluene solution A> Methyl methacrylate 20 parts by mass Methacrylic acid 5 parts by mass 2,2'-azobisisobutyronitrile 0.05 parts by mass Toluene 75 parts by mass

Polymethyl methacrylate hollow fiber (manufactured by Mitsubishi Rayon Co., Ltd., outer diameter 300 μm, inner diameter 200 μm) is 25
The bundle was bundled and one end was fixed with urethane resin so that the hollow portion of the hollow fiber was opened. In the reaction vessel, an ethyl acetate solution B having the following composition was filled into the hollow fibers of this block by suction. Thereafter, the pressure in the reaction vessel was slightly reduced from normal pressure, and a part of the ethyl acetate solution B in the hollow portion was released. The pressure inside the reaction vessel was returned to normal pressure again, and the block was dried overnight in a vacuum dryer to remove ethyl acetate.

<Ethyl acetate solution B> Methyl methacrylate-methacrylic acid copolymer 30 parts by mass 70 parts by mass of ethyl acetate

Example 2 An ethyl acetate solution C having the following composition was prepared, and the inside of the hollow fiber was treated in the same manner as in Example 1.

<Ethyl acetate solution C> 5 parts by mass of methyl methacrylate-methacrylic acid copolymer 95 parts by mass of ethyl acetate

Example 3 Methyl methacrylate-2-
Synthesis of hydroxyethyl methacrylate copolymer Solution D having the following composition was prepared, and methyl methacrylate-2-hydroxyethyl methacrylate copolymer used for the treatment was synthesized. After polymerizing the toluene solution D in a reaction vessel at 70 ° C. for 6 hours under a nitrogen atmosphere, the solution diluted three times with methyl ethyl ketone is poured into seven times the amount of hexane to give methyl methacrylate-2.
-Hydroxyethyl methacrylate copolymer (methyl methacrylate / 2-hydroxyethyl methacrylate = 7
8 mol% / 22 mol%).

<Toluene solution D> Methyl methacrylate 18 parts by mass 2-hydroxyethyl methacrylate 7 parts by mass 2,2'-azobisisobutyronitrile 0.05 parts by mass Toluene 75 parts by mass

Polymethyl methacrylate hollow fiber (produced by Mitsubishi Rayon Co., Ltd., outer diameter 300 μm, inner diameter 200 μm)
The bundle was bundled and one end was fixed with urethane resin so that the hollow portion of the hollow fiber was opened. In a reaction vessel, an ethyl acetate solution E having the following composition was filled into the hollow fibers of this block by suction. Thereafter, the pressure in the reaction vessel was slightly reduced from normal pressure, and a part of the ethyl acetate solution E in the hollow portion was discharged. The pressure inside the reaction vessel was returned to normal pressure again, and the block was dried overnight in a vacuum dryer to remove ethyl acetate.

<Ethyl acetate solution E>-30 parts by mass of methyl methacrylate-2-hydroxyethyl methacrylate copolymer-70 parts by mass of ethyl acetate

Example 4 An ethyl acetate solution F having the following composition was prepared, and the inside of the hollow fiber was treated in the same manner as in Example 1.

<Ethyl acetate solution F> 5 parts by mass of methyl methacrylate-2-hydroxyethyl methacrylate copolymer 95 parts by mass of ethyl acetate

Example 5 The effects of the treatment were confirmed using the blocks treated in the hollow fibers in Examples 1 to 4.
After polymerization of acrylamide gel was performed in each hollow fiber by the following method, the block was sliced in a direction perpendicular to the hollow fiber axis to obtain a thin piece having a thickness of about 750 μm. The flakes are placed in water and incubated at 38 ° C overnight and 50 ° C for 1 hour.
Shake time. After shaking, the slices were observed, and it was confirmed that all 25 hollow fibers were filled with acrylamide gel.

<Polymerization of Acrylamide Gel> An aqueous solution G having the following composition was prepared and filled into the hollow fibers of the blocks prepared in Examples 1 to 4 by suction in a reaction vessel. After filling with aqueous solution, under nitrogen atmosphere 7
Polymerization was carried out at 0 ° C. for 3 hours.

<Aqueous solution G> Acrylamide 9 parts by mass N, N'-methylenebisacrylamide 1 part by mass 2,2'-azobis (2-methylpropionamidine) dihydrochloride (V-50) 0.1 part by mass Water 90 parts by mass Department

Comparative Example 1 Hollow fiber made of polymethyl methacrylate (manufactured by Mitsubishi Rayon Co., Ltd., outer diameter 300 μm, inner diameter 2
(00 μm) were bundled, and one end was hardened with urethane resin so that the hollow portion of the hollow fiber was opened.

Comparative Example 2 <Synthesis of Polystyrene> A toluene solution H having the following composition was prepared, and polystyrene used for the treatment was synthesized. The toluene solution H was polymerized in a reaction vessel at 70 ° C. for 6 hours under a nitrogen atmosphere, and then poured into 5 times the amount of methanol to obtain polystyrene.

<Toluene solution H> Styrene 30 parts by mass 2,2'-azobisisobutyronitrile 0.08 parts by mass Toluene 70 parts by mass

Polymethyl methacrylate hollow fiber (manufactured by Mitsubishi Rayon Co., Ltd., outer diameter 300 μm, inner diameter 200 μm)
The bundle was bundled and one end was fixed with urethane resin so that the hollow portion of the hollow fiber was opened. In a reaction vessel, an ethyl acetate solution I having the following composition was filled into the hollow fibers of this block by suction. Thereafter, the pressure in the reaction vessel was slightly reduced from normal pressure, and a part of the ethyl acetate solution I in the hollow portion was released. The pressure inside the reaction vessel was returned to normal pressure again, and the block was dried overnight in a vacuum dryer to remove ethyl acetate.

<Ethyl acetate solution I> Polystyrene 30 parts by mass Ethyl acetate 70 parts by mass

Comparative Example 3 Using the block prepared in Comparative Example 1, acrylamide was filled in the same manner as in Example 5, and the effect of the treatment was observed. As a result, when slices were obtained by slicing the block, gel loss was observed in 4 of the 25 hollow fibers. After shaking the flakes, a total of 12
Gel loss was observed in the hollow fibers of the book.

Comparative Example 4 Using the block prepared in Comparative Example 2, the effect of the treatment was observed in the same manner as in Example 5. When slicing the blocks to obtain flakes, gel loss was observed in 7 of the 25 hollow fibers. After shaking the flakes, gel loss was observed in a total of 18 hollow fibers.

[0043]

According to the present invention, there is provided a hollow fiber having a hollow portion filled with a gel. Further, the gel inside the hollow fiber filled according to the present invention is hardly removed by being physically fixed to the inner wall portion of the hollow fiber, and the hollow fiber filled with the gel thus produced can be subjected to capillary electrophoresis or DN.
It can be used for manufacturing microarrays for A analysis. In particular, in capillary electrophoresis, the interface between the gel and the inner wall of the capillary is firmly bonded, and there is no short path on the inner wall of the electrophoretic solute such as DNA, and it is expected to perform uniform band-formed electrophoresis. You.

Claims (10)

[Claims] 1. A method for introducing a functional group into the inner wall of a hollow fiber, the method comprising treating a polymer having a hydrophilic group on the inner wall of the hollow fiber. 2. The method for introducing a functional group into the inner wall portion of a hollow fiber according to claim 1, wherein the method for attaching comprises the following steps. (1) a first step of dissolving a polymer having a hydrophilic group in a solvent in advance; (2) a second step of adhering the solution of (1) to the inner wall of the hollow fiber; and (3) a hollow fiber hollow section. A third step of removing the solution and the solvent of the inner wall portion. 3. The method for introducing a functional group into the inner wall of a hollow fiber according to claim 1, wherein the polymer having a hydrophilic group has an anionic group. 4. An anion group comprising a carboxylic acid group, a hydroxyl group,
The method for introducing a functional group according to claim 3, wherein the functional group is at least one selected from the group consisting of sulfonic acid groups. 5. The method for introducing a functional group into the inner wall of a hollow fiber according to claim 1, wherein the polymer having a hydrophilic group has a cationic group. 6. The method according to claim 5, wherein the cationic group is an amino group. 7. The method for introducing a functional group into the inner wall of a hollow fiber according to claim 1, wherein the polymer having a hydrophilic group is a nonion. 8. The method for introducing a functional group into the inner wall of a hollow fiber according to claim 1, wherein the hollow fiber is a non-porous hollow fiber. 9. A gel-forming polymerizable monomer is added to the hollow portion of a hollow fiber in which the polymer having a hydrophilic group obtained by the method according to any one of claims 1 to 8 adheres to the inner wall portion of the hollow fiber. A method of filling a hollow portion of a hollow fiber with a gel, comprising filling and polymerizing the monomer. 10. The gel filling method according to claim 9, wherein the gel-forming polymerizable monomer is a monomer containing acrylamide as a main component.