Reagents for enzyme immunoassay for apolipoprotein B

Claim 1

Reagents for enzyme immunoassay for apolipoprotein B comprising the following components to be used together to perform the immunoassay: human apolipoprotein B as as standard antigen, peroxidase-labeled Fab' specific to apolipoprotein B as an antibody, polystyrene balls coated with IgG which will join with apolipoprotein B by a antigen-antibody reaction, a coloring or fluorescent reagent selected from the group consisting of 3,3',5,5'-tetramethylbenzidine, o-phenylenediamine, 5-aminosalicylate, o-phenyldiamine dihydrochloride, 2,2'-azinodi-(3-ethylbenzthiazoline)-6'-sulfonic acid, thyramine and p-hydroxyphenyl propionate, hydrogen peroxide as a reaction substrate, a reaction stopper, and buffer solutions.

Claim 4

A method for the enzyme immunoassay of apolipoprotein B comprising: (a) coating polystyrene particles with serum immuunoglobulin G (IgG) raised against apolipoprotein B; (b) immersing one or more of said coated polystyrene particles in a serum or saline solution containing apolipoprotein B and a buffer solution containing Fab' specific to apolipoprotein B which Fab' is chemically bound to an peroxidase; (c) transferring said polystyrene particles to a solution containing hydrogen peroxide and, optionally, a reagent selected from the group consisting of 3,3',5,5'-tetramethylbenzidine, o-phenylenediamine, 5-aminosalicylate, o-phenyldiamine dihydrochloride, 2,2'-azinodi-(3-ethylbenzthiazoline)-6' sulfonic acid thyramine and p-hydroxyphenyl propionate; (d) chemically stopping the enzymatic reaction; and (e) measuring the absorption of light or the fluorescence of a enzymatically changed substrate or of the product of the reaction between the enzymatically changed substrate and the reagent.

Claim 5

A method for the enzyme immunoassay of apolipoprotein B comprising: (a) coating polystyrene particles with serum immuunoglobulin G (IgG) raised against apolipoprotein B; (b) immersing one or more of said coated polystyrene particles in a serum or saline solution containing apolipoprotein B and a buffer solution containing Fab' specific to apolipoprotein B which Fab' is chemically bound to an peroxidase; (c) transferring said polystyrene particles to a solution containing hydrogen peroxide and, optionally, a reagent selected from the group consisting of 3,3',5,5'-tetramethylbenzidine, o-phenylenediamine, 5-aminosalicylate, o-phenyldiamine dihydrochloride, 2,2'-azinodi-(3-ethylbenzthiazoline)-6' sulfonic acid thyramine and p-hydroxyphenyl propionate; (d) chemically stopping the enzymatic reaction; and (e) measuring the absorption of light or the fluorescence of a enzymatically changed substrate or of the product of the reaction between the enzymatically changed substrate and the reagent.

Claim 11

A method for the determination of apolipoprotein B in a serum sample, comprising: (a) preparing a saline buffer containing bovine serum albumin; (b) mixing the apolipoprotein B standard, one or more IgG-coated polystyrene balls and peroxidase-labeled Fab' with buffer; (c) incubating the mixture; (d) separating the polystyrene balls from the solution and washing the polystyrene balls in buffer; (e) incubating the polystyrene balls in a solution containing hydrogen peroxide and a reagent which is selected from the group consisting of 3,3',5,5'-tetramethylbenzidine, o-phenylenediamine, 5-aminosalicylate, o-phenyldiamine dihydrochloride, 2,2'-azinodi-(3-ethylbenzthiazoline)-6'-sulfonic acid, o-dianisidine, thyramine and p-hydroxyphenyl propionate; (f) stopping the reaction; (g) measuring the absorbance of light or the fluorescence of the stopped solution; (h) repeating steps (b) through (g) with a different concentration of apolipoprotein B; (i) performing steps (b) through (g) with a sample containing blood serum, and (j) calculating the amount of apolipoprotein B in the serum sample.

Claim 12

A method for the determination of apolipoprotein B in a serum sample, comprising: (a) preparing a saline buffer containing bovine serum albumin; (b) mixing the apolipoprotein B standard, one or more IgG-coated polystyrene balls and peroxidase-labeled Fab' with buffer; (c) incubating the mixture; (d) separating the polystyrene balls from the solution and washing the polystyrene balls in buffer; (e) incubating the polystyrene balls in a solution containing hydrogen peroxide and a reagent which is selected from the group consisting of 3,3',5,5'-tetramethylbenzidine, o-phenylenediamine, 5-aminosalicylate, o-phenyldiamine dihydrochloride, 2,2'-azinodi-(3-ethylbenzthiazoline)-6'-sulfonic acid, o-dianisidine, thyramine and p-hydroxyphenyl propionate; (f) stopping the reaction; (g) measuring the absorbance of light or the fluorescence of the stopped solution; (h) repeating steps (b) through (g) with a different concentration of apolipoprotein B; (i) performing steps (b) through (g) with a sample containing blood serum, and (j) calculating the amount of apolipoprotein B in the serum sample.