Hashida S, Ishikawa S, Nishikata I, Hashinaka K, Oka S, Ishikawa E
Department of Biochemistry, Miyazaki Medical College, Kiyotake, Japan.
J Clin Lab Anal. 1998;12(6):329-36. doi: 10.1002/(sici)1098-2825(1998)12:6<329::aid-jcla1>3.0.co;2-h.
The immune complex transfer enzyme immunoassay for antibody IgM to HIV-1 p17 antigen is described. Serum samples containing antibody IgM to HIV-1 p17 antigen were incubated simultaneously with 2,4-dinitrophenyl-bovine serum albumin-recombinant p17 (rp17) conjugate and rp17-beta-D-galactosidase conjugate, and the immune complex formed comprising the three components was trapped onto colored polystyrene beads coated with affinity-purified (anti-2,4-dinitrophenyl group) IgG. Subsequently, the immune complex was transferred to white polystyrene beads coated with monoclonal mouse (antihuman IgM) IgG in the presence of excess of epsilonN-2,4-dinitrophenyl-L-lysine. The signal for antibody IgM to p17 antigen was the fluorescence intensity by fluorometric assay of beta-D-galactosidase activity bound to the white polystyrene beads. The periods of time required for the formation, trapping, and transferring of the immune complex comprising the three components were more than 4 hr, 2 hr, and 3 hr, respectively. The immunoassay developed was shown to be specific by inhibition of transferring the immune complex in the presence of excess of nonspecific IgM but not IgG. Signals for antibody IgM to p17 antigen with serum samples of HIV-1 seroconversion serum panels,--that is, with serum samples in early stages of the infection--tended to be higher than those with serum samples from HIV-1 asymptomatic carriers probably long after the infection and patients with ARC and AIDS. In contrast, signals for antibody IgG to p17 antigen with serum samples of HIV-1 seroconversion serum panels tended to be higher than signals for antibody IgM to p17 antigen but were much lower than signals for antibody IgG to p17 antigen with serum samples from HIV-1 asymptomatic carriers and patients with ARC and AIDS.
本文描述了用于检测抗HIV-1 p17抗原抗体IgM的免疫复合物转移酶免疫测定法。将含有抗HIV-1 p17抗原抗体IgM的血清样本与2,4-二硝基苯基-牛血清白蛋白-重组p17（rp17）偶联物和rp17-β-D-半乳糖苷酶偶联物同时孵育，形成的包含这三种成分的免疫复合物被捕获到涂有亲和纯化的（抗2,4-二硝基苯基基团）IgG的彩色聚苯乙烯珠上。随后，在过量的εN-2,4-二硝基苯基-L-赖氨酸存在下，将免疫复合物转移到涂有单克隆小鼠（抗人IgM）IgG的白色聚苯乙烯珠上。抗p17抗原抗体IgM的信号是通过对结合在白色聚苯乙烯珠上的β-D-半乳糖苷酶活性进行荧光测定得到的荧光强度。形成、捕获和转移包含这三种成分的免疫复合物所需的时间分别超过4小时、2小时和3小时。所开发的免疫测定法通过在过量非特异性IgM而非IgG存在下抑制免疫复合物的转移而显示出特异性。HIV-1血清转化血清组血清样本（即感染早期的血清样本）中抗p17抗原抗体IgM的信号往往高于HIV-1无症状携带者（可能在感染后很长时间）以及患有艾滋病相关综合征（ARC）和艾滋病患者的血清样本中的信号。相比之下，HIV-1血清转化血清组血清样本中抗p17抗原抗体IgG的信号往往高于抗p17抗原抗体IgM的信号，但远低于HIV-1无症状携带者以及患有ARC和艾滋病患者血清样本中抗p17抗原抗体IgG的信号。
