多种单抗联合检测HIV抗原

目的 建立多种单抗联合早期检测HIV抗原的夹心ELISA方法.方法 以SAS盐析沉淀法和亲和层析法纯化抗HIV-1 p24、gp41、gp120及抗HIV-2 gp36的腹水型单克隆抗体(McAb),用高碘酸钠法将纯化的McAb以HRP进行标记.建立针对单个抗原的双抗体夹心ELISA法,对其灵敏度及特异性进行检测.将筛选得到的4株捕获McAb按比例混合作为捕获抗体,4株酶标McAb按比例混合作为检测抗体,建立多种单抗联合检测HIV抗原的夹心ELISA方法,检测混合HIV抗原.结果 按确定的最优反应条件建立的多种McAb联合夹心ELISA方法,检测到的最高稀释度的HIV混合抗原中各抗原的终浓度分别为:重组HIV-1 p24:0.625 pg/ml,gp41:6.25 ng/ml,gp120:6.25 ng/ml;HIV-2 gp36:9.25 ng/ml.结论 建立了具有高度敏感性的鸡尾酒式多种单抗联合检测HIV抗原的夹心ELISA法,为早期榆测HIV抗原提供了新的思路,为后续的研究奠定了一定基础.

Detection of HIV antigens by mixed several monoclonal antibodies

Objective To establish a sandwich ELISA for early detection of HIV antigens using a mixture of monoclonal antibodies (McAb). Methods The ascites McAbs (anti-HIV-1 p24, anti-HIV-1 gp41, anti-HIV-1 gp120 and anti-HIV-2 gp36) were purified by the SAS and the affinity chromatography,and then were labeled with HRP by sodium metaperiodate. The establishing of sandwich ELISA for detecting the single HIV antigen and the tests of specificity and sensitivity of these systems were performed in advance.A proper ratio mixture of four screened McAbs was used as the capture antibody and a proper ratio mixture of four labeled antibodies was used as the detecting antibody. The method of using sandwich ELISA to detect HIV antigens was set up with these McAbs. Results The sensitivity of this method detecting HIV antigens are:0.625 pg/ml HIV-1 p24, 6.25 ng/ml HIV-I gp41,6.25 ng/ml HIV-I gp120 and 9.25 ng/mi HIV-2 gp36 in mixed HIV antigens. Conclusion The method of using several McAbs mixture in sandwich ELISA detecting HIV antigens was established an excellent sensitivity, which provides a novel idea for early detec-ting the HIV antigen.