猪骨骼肌肌钙蛋白T单克隆抗体的制备及鉴定

为制备猪骨骼肌肌钙蛋白T单克隆抗体，利用生物化学方法提取猪骨骼肌肌钙蛋白并免疫小鼠，通过细胞融合制备稳定分泌单克隆抗体的杂交瘤细胞株，采取体内诱生腹水法批量制备单克隆抗体，并对其免疫学特征进行鉴定。结果表明：最终筛选获得1 株杂交瘤细胞株，将其命名为2E8，其分泌的抗体为免疫球蛋白G 2a亚型，效价为1∶4.1×105，亲和力常数为5.14×106 L/mol；采用间接酶联免疫吸附测定检测单克隆抗体2E8特异性，结果表明，单克隆抗体2E8与鸡、鸭检测原的交叉反应率均为25%；与牛、羊检测原的交叉反应率分别为1.7%、6.3%；免疫印迹检测结果显示，单克隆抗体2E8与猪检测原反应条带清晰，与羊、鸡、鸭检测原反应条带轻微，与牛检测原几乎不发生反应。

Preparation and Characterization of Monoclonal Antibody against Porcine Skeletal Muscle Troponin T

In order to prepare monoclonal antibodies against porcine skeletal muscle troponin T (sTnT), porcine skeletal muscle troponins were extracted by biochemical methods and used to immunize mice. By means of cell fusion, we acquired hybridoma cell strains that can secrete monoclonal antibodies (mAbs) stably. Ascites was induced in vivo to further prepare and characterize monoclonal antibodies against sTnT. Finally, we screened out one hybridoma cell strain that secretes mAb stably and named it 2E8. The monoclonal antibody 2E8 belonged to the IgG2a subtype, whose titer and affinity constant were 1 : 4.1 × 105 and 5.14 × 106 L/mol, respectively. The specificity of 2E8 was evaluated by indirect enzyme-linked immunosorbent assay (iELISA). The cross-reactivity with chicken sTnT and duck sTnT were both 25%. The cross-reactivity with bovine sTnT and goat sTnT were 1.7% and 6.3% respectively. Immunoblotting data showed that the monoclonal antibody 2E8 exhibited a clear band after reaction with the porcine antigen, but only a slight band after reaction with the goat, chicken, or duck antigen. In addition, the monoclonal antibody hardly reacted with the bovine antigen.