Open AccessDetection of antibody IgG to HIV‐1 in urine by ultrasensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) using recombinant p24 as antigen for diagnosis of HIV‐1 infection
Author(s) -
Hashida Seiichi,
Hashinaka Kazuya,
Hirota Kouichi,
Saitoh Atsushi,
Nakata Atsuo,
Shinagawa Hideo,
Oka Shinichi,
Shimada Kaoru,
Mimaya JunIchi,
Matsushita Shuzo,
Ishikawa Eiji
Publication year - 1994
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.1860080206
Subject(s) - immunoassay , antibody , recombinant dna , antigen , immune complex , microbiology and biotechnology , chemistry , affinity chromatography , bovine serum albumin , immunoglobulin g , conjugate , protein g , urine , virology , enzyme , chromatography , biology , immunology , biochemistry , gene , mathematical analysis , mathematics
Abstract Anti‐HIV‐1 IgG in urine was detected by an ultrasensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) using recombinant p24 gag protein (p24) of HIV‐1 as antigen and β‐D‐galactosidase from Escherichia coli as label. Anti‐HIV‐1 IgG in urine was reacted simultaneously with 2, 4‐dinitrophenyl‐bovine serum albumin‐recombinant p24 conjugate and recombinant p24‐β‐D‐galactosidase conjugate. 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 ϵN‐2, 4‐dinitrophenyl‐L‐lysine, and transferred to polystyrene balls coated with affinity‐purified (anti‐human IgG γ‐chain) IgG. Bound γ‐D‐galactosidase activity was assayed by fluorometry. This assay was at least 3, 000‐fold more sensitive than conventional methods. The lowest signal among 49 asymptomatic carriers was 3.1‐fold higher than the highest nonspecific signal among 100 seronegative subjects. The sensitivity and specificity were both 100%. The positivity could be confirmed by preincubation of urine samples with excess of the antigen. Thus, this assay would be a powerful tool for detecting IgG antibody to HIV‐1 in urine. © 1994 Wiley‐Liss, Inc.