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目的利用杂交瘤技术制备分泌抗重组旋毛虫副肌球蛋白N端抗原(rTsP3)的单克隆抗体(McAb)并进行鉴定。方法以rTsP3免疫BALB/c小鼠,取其脾细胞与小鼠骨髓瘤SP2/0细胞融合,筛选分泌高滴度McAb杂交瘤细胞株,制备腹水并进行纯化,采用间接ELISA法测定培养细胞上清液及腹水中的McAb滴度、相对亲和力及抗体亚类,Western blot法鉴定抗体对抗原识别的特异性。结果获得了2株稳定分泌抗旋毛虫rTsP3的McAb杂交瘤细胞株,分泌的McAb分别为IgG2b亚类κ型和IgG1亚类κ型,亲和力常数分别为8.98×108mol/L和9.7×108mol/L,Western blot显示2株单抗均能识别旋毛虫成虫匀浆蛋白、rTsP3及重组副肌球蛋白(rTsPmy)。结论成功制备了抗旋毛虫rTsP3单克隆抗体,该单抗能识别旋毛虫副肌球蛋白抗原。
OBJECTIVE: To clone and identify McAbs secreting anti-Trichinella spiralis paramyosin N-terminal antigen (rTsP3) by hybridoma technology. Methods BALB / c mice were immunized with rTsP3 and their spleen cells were fused with mouse myeloma SP2 / 0 cells. The McAbs secreting high titer McAbs were screened and secreted. The ascites was prepared and purified. McAb titers, relative affinities and antibody subclasses in serum and ascites, Western blot were used to identify the specificity of the antibody for antigen recognition. Results Two McAbs hybridomas secreting anti-Trichinella rTsP3 were obtained. McAbs secreted by the McAbs were IgG2b subclass κ and IgG1 subclass κ with affinity constants of 8.98 × 108mol / L and 9.7 × 108mol / L, respectively Western blot showed that the two monoclonal antibodies could recognize Trichinella spiralis adult homogenate, rTsP3 and recombinant paramyosin (rTsPmy). Conclusion The anti-Trichinella rTsP3 monoclonal antibody was successfully prepared, which can recognize Trichinella spiralis paramyosin antigen.