抗HBeAg不同位点的单克隆抗体杂交瘤细胞株的制备及其应用

Establishment and application of hybridoma cell lines secreting monoclonal antibodies against two distinct antigenic determinants of hepatitis B e antigen

目的建立检测人乙型肝炎病毒e抗原(HBeAg)的双单抗夹心酶联免疫吸附试验(ELISA)检测试剂。方法用基因工程HBeAg免疫Balb/c小鼠,采用杂交瘤技术建立能稳定分泌针对抗原不同位点单抗的杂交瘤细胞株。将构象型位点单抗和线性位点单抗配对做夹心ELISA,选择最匹配的一对建立双单抗夹心HBeAg检测试剂,并与人抗HBeAg多抗包板、单抗标酶的HBeAg检测试剂盒做对比试验。结果建立了89株能稳定分泌小鼠抗HBeAg的杂交瘤细胞株,其中63株单抗针对构象型位点,26株单抗针对线性位点。挑选出最匹配的7E6和e1构建成检测HBeAg的双单抗夹心ELISA检测试剂。与人抗HBeAg多抗包被、单抗酶标检测试剂对比,双单抗试剂具有更高的灵敏度,而其他指标基本一致。结论我们用不同位点的HBe单克隆抗体建立了高灵敏度、高特异性的有实用价值的HBeAg双单抗夹心ELISA检测试剂。

Objective To establish a two-site immunoassay for hepatitis B e antigen （HBeAg）. Methods Balb/ c mice were immunized with recombinant HBeAg, and their spleen cells were fused with mouse myeloma cells SP 2/0. Cell lines secreting antibodies against two distinct antigenic determinants on HBeAg were obtained. An enzyme-linked immunosorbent assay （ELISA） kit for HBeAg was developed using a matched monoclonal antibody pair, which was com- pared with a commercial kit employing polyclonal and monoclonal antibody. Results 89 hybridoma cell lines secreting antibodies against HBeAg were obtained, among them 63 recognize a linear antigenic determinant and 26 recognize a conformational one. A best matched antibody pair, 7E6 and el were selected and used to develop an ELISA kit for detecting HBeAg. Compared with the commercial kit, the monoclonal-monoclonal sandwich ELISA kit had a higher sensitivity and better specificity. Conclusions The two-site ELISA kit established for detecting HBeAg has a higher sensitivity and better specificity.