脾内注射法免疫小鼠制备抗-HBc单克隆抗体的研究

DEVELOPMENT OF MOUSE ANTI-HBc MONOCLONAL ANTIBODIES BY INTRASPLENIC IMMUNIZATION

用基因工程菌产乙型肝炎核心抗原(HBcAg)对 Balb/c 小鼠脾内多点注射法免疫,将该鼠的脾淋巴细胞与鼠骨髓瘤细胞在 PEG 作用下融合,用固相放射免疫分析法(SPRIA)筛选,成功地获得了2株抗-HBc 阳性的杂交瘤细胞株。通过中和试验和交叉鉴定,2株抗-HBc 单克隆抗体只能与 HBcAg 起反应,而不能与 HBsAg 和 HBeAg 反应。经多次亚克隆后,所得腹水中的抗-HBc 滴度达1:64000～1:28000。这2株单克隆抗体做包被抗体检测 HBcAg 和(125)~Ⅰ标记查患者血清标本时,所测得的结果与人抗-HBc 多克隆抗体做包被或(125)~Ⅰ标记所测得的结果相一致。2株杂交瘤细胞连续传代3个多月,冻存复苏后仍能稳定地分泌特异的抗-HBc 单克隆抗体。

Balb/c mice were immunized with HBcAg purified from engineered E coli strain by intrasplenic injections.The spleen cells were fused with mouse myeloma cells in the prese- nce of PEG,and then two clones of hybridoma cells secreting anti-HBc were acquired.The monoclonal antibodies were found to react only with HBcAg and were comparable to human polyclonal antibodies to HBcAg when they were used to detect HBcAg or anti-HBc antibo- dies in sera specimens from patients.The titer of the monoclonal antibodies in mouse ascites could reach 1:64 000-1:128 000 by SPRIA after repeated subcloning.The two clones of hybridoma cells can stably produce the specific anti-HBc monoclonal antibodies during conti- nuously cultivation for more than 3 months or after frozen and thawed.