抗乙肝病毒前S(2)蛋白单克隆抗体杂交瘤细胞株的建立及初步应用

用聚合人血清白蛋白亲和层析柱纯化的含前S(2)-HBsAg,脾内接种Balb/c小鼠,取其脾细胞与SP2/0-Ag骨髓瘤细胞融和,以4种酶联法进行筛选,并经多次有限稀释亚克隆培养,最终选出3株分泌抗前S(2)单克隆抗体的细胞株。经多次传代,结果稳定。接种Balb/c小鼠腹腔后,腹水均为抗前S(2)阳性,效阶达8万。免疫球蛋白为鼠IgG1型。用其初步纯化的IgG酶标记物,以双抗体夹心法检测了乙肝病人血清中前S(2)蛋白,结果较满意,认为可在临床上推广使用。

THE ESTABLISHMENT OF HYBRIDOMA CELLS PRODUCING MONOCLONAL ANTIBODY TO PRE- S (2) EPITOPE OF HBV AND ITS PRELIMINARY CLINICAL USE

HBsAg containing pre-s (2), purified by affinity chromatography with PHSA coupled to CNBr-activated sepharose 4B, was used as immunogen to immunize in vivo spleen of BALB/C female mice. The spleen cells from immunized mice were fused with mouse myeloma cells (SP2/0-Ag). Clones producing the antibody were screened by serial enzyme immunosorbent assay, and subclone cultures were isolated by limiting dilution. Finally, three clones, which were positive for anti-pre-s (2) antibody, were selected. The clones were still stable by several passages. They were injected i. p. to prime-treated BALB/C mice to produce ascitic fluid. The liters of anti-pre-s (2) antibodies in the ascitic fluid were above 105.Isotype analysis revealed that all immunoglobulins were of IgGl subclass of mouse. The purified McAb for pre-s (2) conjugated with horseradish peroxidase were used to develop a sandwich-type assay (ELISA). Using the assay, pre-s (2), the receptor for PHSA,was detected in serum of the patients with HBV infection. The assay method was more sensitive, specific, reproducible and reliable. The assay may be practical in the routine work of hepatitis B immunodiagnosis, and worth further clinical trial.