Detection of antibody IgG to HIV-1 in urine by sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) using recombinant proteins as antigens for diagnosis of HIV-1 infection.
For diagnosis of HIV-1 infection, attempts were made to detect anti-HIV-1 IgG in urine by sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) using recombinant reverse transcriptase (RT) and p17 as antigens. Anti-HIV-1 IgG in urine was reacted simultaneously with 2,4-dinitrophenyl-bovine serum albumin-recombinant protein conjugate and recombinant protein-enzyme conjugate. The enzymes used as labels were horseradish peroxidase for RT and Escherichia coli beta-D-galactosidase for p17. The complex formed, consisting of the three components, was trapped onto polystyrene balls coated with affinity-purified (anti-2,4-dinitrophenyl group) IgG, eluted with epsilon N-2,4-dinitrophenyl-L-lysine and transferred to polystyrene balls coated with affinity-purified (anti-human IgG gamma-chain) IgG. Finally, bound enzyme activity was assayed by fluorometry. Urine samples were collected from 100 seronegative subjects and 70 seropositive subjects. The sensitivity and specificity were both 100% with unconcentrated urine samples. The positivity was confirmed by preincubation of urine samples with excess of the antigens. The positivity and negativity with one of the two antigens could be confirmed with the other antigen. The positivity with low signals could be confirmed by concentration of urine samples. Detection of anti-HIV-1 IgG in urine by the immune complex transfer enzyme immunoassay using different antigens would make diagnosis of HIV-1 infection possible.
Hashida
S
S
Department of Biochemistry, Medical College of Miyazaki, Japan.
Hirota
K
K
Hashinaka
K
K
Saitoh
A
A
Nakata
A
A
Shinagawa
H
H
Oka
S
S
Shimada
K
K
Mimaya
J
J
Matsushita
S
S
D000293:Adolescent; D000328:Adult; D000368:Aged; D002648:Child; D005069:Evaluation Studies as Topic; D015683:Gene Products, gag / Q000276:immunology; D015483:HIV Antibodies / Q000652:urine*; D015488:HIV Antigens / Q000276:immunology; D015658:HIV Infections / Q000175:diagnosis* / Q000276:immunology / Q000652:urine; D054303:HIV Reverse Transcriptase; D018023:HIV Seronegativity / Q000276:immunology; D006679:HIV Seropositivity / Q000175:diagnosis / Q000276:immunology / Q000652:urine; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D007124:Immunoenzyme Techniques / Q000706:statistics & numerical data; D007074:Immunoglobulin G / Q000652:urine*; D008875:Middle Aged; D012194:RNA-Directed DNA Polymerase / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D012680:Sensitivity and Specificity; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus
D015683:Gene Products, gag; D015483:HIV Antibodies; D015488:HIV Antigens; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; C054858:p17 protein, Human Immunodeficiency Virus Type 1; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase