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目的 研制可同时检测HIV 1、HIV 2抗体及p2 4抗原的胶体金快速诊断试剂。方法 利用重组杆状病毒 昆虫细胞系统进行HIV 1gp4 1及HIV 2gp36抗原的高效表达 ,以免疫亲和层析法纯化抗原。抗 HIVp2 4单克隆抗体杂交瘤细胞株 ,并制备抗 HIVp2 4单克隆抗体。以硝酸基纤维膜为载体 ,以纯化的HIV 1gp4 1、HIV 2gp36抗原及抗p2 4抗体点膜 ,2 0nm胶体金颗粒 抗人IgG和抗 HIVp2 4单克隆抗体进行标记 ,对 33份已知HIV感染者阳性血清及 6份阴性血清进行检测。结果 通过重组杆状病毒 昆虫细胞系统进行HIV 1gp4 1及HIV 2gp36抗原的表达 ,可获取浓度为 2 .0mg L的纯化抗原。从抗p2 4单克隆抗体杂交瘤细胞株中培养上清液 ,通过葡萄球菌蛋白A免疫亲和层析柱可得到 1.5mg L的纯化抗体。利用纯化的抗原抗体进行标记 ,对 39份已知血清进行检测 ,与荷兰Organon公司HIV1+2抗体、p2 4抗原、ELISA诊断试剂同时检测结果进行比较 ,证实有较强的特异性及敏感性。结论 同时检测HIV 1、HIV 2抗体及p2 4抗原快速诊断试剂的问世 ,可为HIV感染的诊断提供一个简便、可靠、敏感的方法。
Objective To develop a colloidal gold rapid diagnostic reagent that can simultaneously detect HIV 1, HIV 2 and p2 4 antigens. Methods High-level expression of HIV 1 gp41 and HIV 2 gp36 antigens was achieved by recombinant baculovirus insect cell system, and the antigen was purified by immunoaffinity chromatography. Anti-HIVp2 4 monoclonal antibody hybridoma cell line and preparation of anti-HIVp2 4 monoclonal antibody. Using nitrocellulose membrane as a carrier, purified HIV 1 gp41, HIV 2 gp36 antigen and anti-p2 4 antibody spots, 20 nm colloidal gold particle anti-human IgG and anti-HIVp2 4 monoclonal antibody were labeled. 33 samples of known HIV Infected positive sera and 6 negative sera were tested. Results The purified antigen was obtained at a concentration of 2.0 mg L by recombinant baculovirus insect cell system for the expression of HIV 1 gp41 and HIV 2 gp36 antigens. From the anti-p2 4 monoclonal antibody hybridoma cell culture supernatant, staphylococcal protein A immunoaffinity chromatography can be obtained 1.5mg L purified antibody. Using purified antigen and antibody for labeling, 39 samples of known serum were tested and compared with the results of simultaneous detection of Organon HIV1 + 2 antibody, p2 4 antigen and ELISA diagnostic reagent from Holland Company. The results confirmed that the method has high specificity and sensitivity. Conclusion Simultaneous detection of HIV 1, HIV 2 antibody and p2 4 antigen rapid diagnostic reagent can provide a simple, reliable and sensitive method for the diagnosis of HIV infection.