鸭骨骼肌肌钙蛋白I单克隆抗体的制备及其特性

提取鸭骨骼肌肌钙蛋白I（skeletal muscle troponin I，sTnI）用于制备单克隆抗体，并对抗体性能进行 评价。将鸭骨骼肌提取后经十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（sodium dodecyl sulfate polyacrylamide gel electropheresis，SDS-PAGE）测定，取凝胶条带免疫小鼠，融合后经筛选获得分泌抗鸭sTnI的杂交瘤细胞株，用 酶联免疫吸附测定法检测抗体效价、亚型及特异性等特性。结果表明：鸭骨骼肌提取物经SDS-PAGE后出现2 条 颜色较深的条带，取分子质量24 kDa条带用于免疫；筛选后获得2 株单克隆抗体，其中3E7特异性较好，效价在 1∶1.0×105以上，为IgG1亚型，亲和常数（Ka）为5.9×105 L/mol；免疫印迹结果显示，抗体3E7能与鸭骨骼肌提取 物反应，与牛、羊、鸡、鱼骨骼肌提取物无明显反应。

Preparation and Characterization of Monoclonal Antibody against Duck Skeletal Muscle Troponin I

A monoclonal antibody (mAb) against duck skeletal muscle troponin I (sTnI) was prepared and its characteristics were evaluated. sTnI was analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE). The protein band was excised from the gel and used to immunize mice. Hybridoma cell strains stably secreting mAb were obtained by fusion. The titer, subtype and speci?city of monoclonal antibodies were analyzed by enzyme linked immunosorbent assay. The SDS-PAGE showed 2 dark bands. The 24 kDa protein was selected for immunization. Two cell lines that secreted monoclonal antibody against sTnI were obtained after screening, with 3E7 being the one with higher speci?city. It belonged to the IgG1 subtype, its titer was 1:1.0 × 105 and its af?nity constant was 5.9 × 105 L/mol. Western blot results showed that 3E7 reacted with duck skeletal muscle extracts from duck, but not bovine, sheep, chicken and ?sh skeletal muscle extracts.