Highly reproducible agglutination immunoassay method and reagents
View Patent ↗The present invention provides an agglutination immunoassay, wherein the agglutination of insoluble carrier particles such as latex are stabilized and uniformized to give good reproducibility, and a reagent therefor. In the agglutination immunoassay which comprises allowing an antigenic substance in a sample to bind to insoluble carrier particles carrying substantially neither antigens nor antibodies thereon, and allowing an antibody or an antibody complex which reacts specifically to the antigenic substance to bind to the antigenic substance to give a selective agglutination of the insoluble carrier particles, a homopolymer prepared by polymerization of a monomer such as 2-methacryloyloxyethyl phosphorylcholine having a phosphorylcholine group and a vinyl group, or a copolymer prepared by polymerization of a monomer having a phosphorylcholine group and a vinyl group, with a monomer having a vinyl group such as n-butyl methacrylate is used.
1. An agglutination immunoassay for determining the presence and/or amount of an antigenic substance in a sample, which comprises
allowing an antigenic substance in a sample to bind to insoluble carrier particles carrying substantially neither antigens nor antibodies thereon, and
allowing an antibody or an antibody complex which reacts specifically to the antigenic substance to bind to the antigenic substance to give a selective agglutination of the insoluble carrier particles in the presence of a compound having a group represented by formula (I),
wherein n is an integer of 1 to 6, and R 1 , R 2 and R 3 are the same or different, and independently represent hydrogen, substituted or unsubstituted alkyl having 1 to 6 carbon atoms, and
measuring the degree of selective agglutination of the insoluble carrier particles, which indicates the presence and/or amount of antigenic substance in the sample.
2. The agglutination immunoassay according to claim 1 , wherein the compound having a group represented by formula (I) is a compound prepared by polymerization of a monomer having a group represented by formula (I).
3. The agglutination immunoassay according to claim 1 , wherein the compound having a group represented by formula (I) is a compound prepared by polymerization of a monomer having a group represented by formula (D with other monomer polymerizable with the monomer having a group represented by formula (I).
4. The agglutination immunoassay according to claim 3 , wherein the other monomer polymerizable with the monomer having a group represented by formula (I) is a monomer having a vinyl group.
5. The agglutination immunoassay according to claim 4 , wherein the monomer having a vinyl group is n-butyl methacrylate.
6. The agglutination immunoassay according to claim 2 , wherein the monomer having a group represented by formula (I) is a monomer having a group represented by formula (I) and a vinyl group.
7. The agglutination immunoassay according to claim 1 , wherein the group represented by formula (I) is a phosphorylcholine group.
8. The agglutination immunoassay according to claim 6 , wherein the monomer having a group represented by formula (I) and a vinyl group is 2-methacryloyloxyethyl phosphorylcholine.
9. The agglutination immunoassay according to claim 1 , wherein the antibody is an anti-hemoglobin A1c monoclonal antibody.
10. The agglutination immunoassay according to claim 1 , wherein the antibody complex comprises an antibody which reacts specifically to the antigenic substance, and a secondary antibody which reacts selectively to the antibody which reacts specifically to the antigenic substance.
11. The agglutination immunoassay according to claim 1 , wherein the insoluble carrier particles are polystyrene latex.
12. A reagent kit for an immunoassay comprising an insoluble carrier particle carrying substantially neither antigens nor antibodies thereon, an antibody that binds to an antigen to be determined in a sample, and a compound having a group represented by formula (I),
wherein n is an integer of 1 to 6, and R 1 , R 2 and R 3 are the same or different, and independently represent hydrogen, substituted or unsubstituted alkyl having 1 to 6 carbon atoms.
13. The reagent kit for an immunoassay according to claim 12 , wherein the compound having a group represented by formula ( 1 ) is a compound prepared by polymerization of a monomer having a group represented by formula (I).
14. The reagent kit for an immunoassay according to claim 12 , wherein the compound having a group represented by formula (D is a compound prepared by polymerization of a monomer having a group represented by formula (I) with other monomer polymerizable with the monomer having a group represented by formula (I).
15. The reagent kit for an immunoassay according to claim 14 , wherein the other monomer polymerizable with the monomer having a group represented by formula (I) is a monomer having a vinyl group.
16. The reagent kit for an immunoassay according to claim 15 , wherein the monomer having a vinyl group is n-butyl methacrylate.
17. The reagent kit for an immunoassay according to claim 13 , wherein the monomer having a group represented by formula (I) is a monomer having a group represented by formula (I) and a vinyl group.
18. The reagent kit for an immunoassay according to claim 12 , wherein the group represented by formula (I) is a phosphorylcholine group.
19. The reagent kit for an immunoassay according to claim 17 , wherein the monomer having a group represented by formula (I) and a vinyl group is 2-methacryloyloxyehtyl phosphorylcholine.
20. The reagent kit for an immunoassay according to claim 12 , wherein the antibody is an anti-hemoglobin A1c monoclonal antibody.
21. The reagent kit for an immunoassay according to claim 12 , wherein the insoluble carrier particles are polystyrene latex.
22. The reagent kit for an immunoassay according to claim 12 , further comprising a secondary antibody that binds to the antibody that binds to the antigen to be determined in the sample.
23. The reagent kit for an immunoassay according to claim 12 , further comprising a buffer.