Novel and sensitive enzyme immunoassay (immune-complex-transfer enzyme immunoassay) for antihuman t cell leukemia virus type 1 igg in human serum using recombinant gag-env hybrid protein as antigen

✍ Scribed by Hiroaki Kohno; Takeyuki Kohno; Eiji Ishikawa; Iwane Sakoda

• Publisher: John Wiley and Sons
• Year: 1990
• Tongue: English
• Weight: 736 KB
• Volume: 4
• Category: Article
• ISSN: 0887-8013
• DOI: 10.1002/jcla.1860040508

✦ Synopsis

A novel and sensitive enzyme immunoassay (immune-complex-transfer enzyme immunoassay) for antihuman T cell leukemia virustype 1 IgG (anti-HTLV-1 IgG) in human serum using recombinant gag( 14-1 39)-env-(1 97-295) hybrid protein is described. Anti-HTLV-1 IgG in test serum was reacted with dinitrophenyl biotinyl bovine serum albuminrecombinant gag-env hybrid protein conjugate. The complex formed was trapped onto polystyrene balls coated with affinity-purified antidinitrophenyl group IgG. After washing to eliminate nonspecific IgG in the test serum, the complex was eluted from the polystyrene balls with dinitrophenyl-L-lysine and transferred to polystyrene balls coated with streptavidin. After washing, anti-HTLV-1 IgG in the complex trapped onto the streptavidin-coated polystyrene balls was reacted with antihuman IgG y-chain Fab'-peroxidase conjugate. Peroxidase activity bound to the streptavidincoated polystyrene balls was assayed by fluorometiy. By transfer of the complex, the nonspecific binding of nonspecific human IgG was considerably reduced, and the detection limit of anti-HTLV-1 IgG in serum was lowered 30-300-fold compared with that by Western blotting, gelatin particle agglutination, and the conventional enzyme immunoassay, in which a recombinant gag-env hybrid protein-coated polystyrene ball was incubated with test serum and, after washing, with antihuman IgG ?-chain Fab'-peroxidase conjugate. Usefulness of the immune-complex-transfer enzyme immunoassay was demonstrated using 271 serum samples.