Dry detection reagent containing acrylamide/styrene copolymer particles immobilizing an immunologically active substance

• US 5,656,506 A
• Filed: 06/01/1995
• Issued: 08/12/1997
• Est. Priority Date: 07/13/1990
• Status: Expired due to Term

• Current Assignee: Canon Kabushiki Kaisha (Canon Inc.)
• Original Assignee: Canon Kabushiki Kaisha (Canon Inc.)
• Inventors: Miyazaki, Takeshi, Kawaguchi, Haruma
• Primary Examiner(s): Naff, David M.
• Application Number:
  US08/456,622
• Time in Patent Office: 803 Days
• Field of Search: 435/7.1, 435/180, 435/7.92, 436/531, 436/533, 436/534, 530/815
• US Class Current: 436/534
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form
• CPC Class Codes: G01N 33/545 Synthetic resin
• CPC Class Codes: Y10S 530/815 Carrier is a synthetic polymer

Abstract

A dry detection reagent for detecting an immunologically active substance is prepared containing solid fine polymer particles immobilizing a substance immunologically active for the substance detected. The polymer particles are prepared by one of the following methods: a) ternary copolymerization of a monomer having carboxyl groups, a monomer having amino groups and styrene or its derivatives; b) binary copolymerization of a monomer having carboxyl groups and styrene or its derivatives, and reacting part of the carboxyl groups with a bifunctional amine such as ethylene diamine to provide amino groups; or c) copolymerizing styrene or its derivatives with a monomer having amide groups, and converting part of the amide groups to amino groups and carboxyl groups with the Hofmann reaction, or hydrolyzing part of the amide groups to form carboxyl groups. Preferably, styrene and acrylamide having amide groups are copolymerized and part of the amide groups are hydrolyzed to carboxyl groups or part of the amide groups are converted to carboxyl groups and amino groups by the Hofmann reaction. The immunologically active substance used for detecting is immobilized on the particles and the particles are dried to produce the dry reagent. To carry out detection, the dry reagent is dispersed in a liquid, the liquid is mixed with a sample containing the substance to be detected which becomes bound to the substance immobilized on the particles and the bound substance is optically detected. The dry reagent is stable during storage and has excellent re-dispersibility in a liquid.

Claims

1. A process for detecting a first substance being immunologically active in a sample by using a diagnostic reagent, comprising the steps of;

(a) preparing a dry diagnostic reagent including a second substance immunologically active with the first substance by the steps of;

(i) providing solid fine particles consisting essentially of a copolymer consisting of styrene and acrylamide having amide groups as copolymerized monomers;

(ii) incorporating carboxyl groups into the particles by hydrolyzing a part of the amide groups contained by the acrylamide of the copolymer, to obtain particles having carboxyl groups and amide groups in a ratio of 0.01 to 1,000 carboxyl groups per one amide group;

(iii) combining the second substance with the particles having the carboxyl groups and the amide groups whereby the second substance becomes bound to the particles; and

(iv) drying the particles containing the bound second substance to prepare the dry diagnostic reagent;

(b) dispersing the dry diagnostic reagent in a liquid;

(c) mixing the liquid in which the diagnostic reagent is dispersed with a sample containing the first substance to bind the first substance to the second substance; and

(d) optically detecting the first substance bound to the second substance contained by the diagnostic reagent.

2. The process according to claim 1, wherein the ratio of the carboxyl groups to the amide groups is 0.5 to 100 carboxyl groups per one amide group.

3. A process for detecting a first substance which is immunologically active in a sample, comprising the steps of;

(a) preparing a dry diagnostic reagent including a second substance immunologically active with the first substance by the steps of;

(i) providing solid fine particles consisting essentially of a copolymer consisting of styrene and acrylamide having amide groups as copolymerized monomers;

(ii) incorporating carboxyl carboxyl groups into the particles by hydrolyzing a part of the side groups contained by the acrylamide of the copolymer, and incorporating amino groups into the particles by converting a portion of the amide groups, contained by the acrylamide of the copolymer to amino groups using the Hofmann reaction, to obtain particles having carboxyl groups and amino groups in a ratio of 0.01 to 100 carboxyl groups per one amino group;

(iii) combining the second substance with the particles having the carboxyl groups and the amino groups whereby the second substance becomes bound to the particles;

(iv) drying the particles containing the bound second substance to prepare the dry diagnostic reagent;

(b) dispersing the dry reagent in a liquid;

(c) mixing the liquid in which the diagnostic reagent is dispersed with a sample containing the first substance to bind the first substance to the second substance; and

(d) optically detecting the first substance bound to the second substance contained by the diagnostic reagent.

4. The process according to claim 3, wherein the ratio of the carboxyl groups to the amino groups is 0.5 to 20 carboxyl groups per one amino group.

5. A dry detection reagent for detecting a first immunologically active substance in a sample comprising:

solid fine particles consisting of a copolymer consisting of styrene and acrylamide as copolymerized monomers containing carboxyl groups and amino groups in a ratio of from 0.01 to 100 carboxyl groups per one amino group, which are attached to a main chain of the copolymer, said solid fine particles having immobilized thereon a second substance which is immunologically active for the first substance to be detected, and said fine particles containing said immobilized second substance being in dry form.

6. A dry detection reagent for detecting a first immunologically active substance in a sample comprising:

solid fine particles consisting of a copolymer consisting of styrene and acrylamide as copolymerized monomers containing carboxyl groups and amide groups in a ratio of from 0.01 to 1,000 carboxyl groups per one amide group, which are attached to a main chain of the copolymer, said solid fine particles having immobilized thereon a second substance which is immunologically active for the first substance to be detected, and said fine particles containing said immobilized second substance being in dry form.