Immune complex transfer enzyme immunoassay for antibody IgG to HIV-1 gp41 antigen using synthetic peptides as antigens.
The immune complex transfer enzyme immunoassay for antibody IgG to HIV-1 gp41 antigen was developed using two synthetic peptides. An aliquot (10 microl) of serum samples from HIV-1 seropositive subjects was incubated simultaneously with 2,4-dinitrophenyl-bovine serum albumin-synthetic HIV-1 gp41 peptide conjugates and synthetic HIV-1 gp41 peptide-beta-D-galactosidase conjugates and subsequently with colored polystyrene beads coated with affinity-purified (anti-2,4-dinitrophenyl group) IgG to trap the immune complexes formed comprising the three components. After washing, the colored polystyrene beads were incubated with white polystyrene beads coated with affinity-purified (anti-human IgGgamma-chain) IgG in the presence of epsilonN-2,4-dinitrophenyl-L-lysine to transfer the immune complexes to the white polystyrene beads. Beta-D-Galactosidase activity bound to the white polystyrene beads was assayed by fluorometry. The formation, trapping and transferring of the immune complexes were completed within 0.5, 0.5 and 1.5 hr, respectively. Since each peptide appeared to react with its own specific antibody IgG, serum samples were tested by the equimolar combination of the two peptides. The lowest signals (fluorescence intensities for bound beta-D-galactosidase activity) for serum samples from HIV-1 asymptomatic carriers, patients with AIDS-related complex and patients with AIDS were 1490-, 2210- and 1460-fold, respectively, higher than the highest signal for serum samples from HIV-1 seronegative subjects. In five seroconversion serum panels, antibody IgG to HIV-1 gp41 antigen was detected as early as antibodies to HIV-1 detected by three currently commercially available methods.
Department of Biochemistry, Miyazaki Medical College, Kiyotake, Japan.
S
S
Hashida
S
S
Ishikawa
K
K
Hashinaka
I
I
Nishikata
S
S
Oka
E
E
Ishikawa
D000163:Acquired Immunodeficiency Syndrome / Q000097:blood; D000328:Adult; D000368:Aged; D000936:Antigen-Antibody Complex / Q000097:blood* / Q000276:immunology; D005260:Female; D015488:HIV Antigens / Q000097:blood* / Q000276:immunology; D015700:HIV Envelope Protein gp41 / Q000097:blood* / Q000737:chemistry / Q000276:immunology; D006679:HIV Seropositivity / Q000097:blood; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000097:blood* / Q000276:immunology; D008297:Male; D008875:Middle Aged; D010455:Peptides / Q000097:blood* / Q000737:chemistry / Q000276:immunology; D013997:Time Factors
D000936:Antigen-Antibody Complex; D015488:HIV Antigens; D015700:HIV Envelope Protein gp41; D007074:Immunoglobulin G; D010455:Peptides