抗人心肌肌凝蛋白轻链Ⅰ单克隆抗体的制备和鉴定

PREPARATION AND CHARACTERIZATION OF MONOCLONAL ANTIBODIES AGAINST HUMAN CARDIAC MYOSIN LIGHT CHAIN

以人心肌肌凝蛋白轻链I免疫Balb/c小鼠,取其脾淋巴细胞与NS-1细胞融合,经过筛选、克隆化和再克隆已获得3-D_3、2-D_9、1-E_(10)和2-E_(10)等4个分泌特异性单克隆抗体的杂交瘤细胞株。经免疫酶标和放免方法测定抗体工作滴度为1:1万。流动式荧光密度计检测NS-1、小鼠脾和杂交瘤细胞DNA的含量,结果表明杂交瘤细胞的DNA含量相当于NS-1和小鼠脾细胞DNA含量之和。杂文瘤细胞传代半年以上稳定地分泌特异性的单克隆抗体。用两个单抗进行了抗原双决定簇的放免測定,并制备了高效价的小鼠腹水。

Four hybridoma cell lines secreted specific monoclonal antibodies were obtained by fusion of NS-1 mouse myeloma cells with spleen cells of Balb/c mouse which was immunized with human cardiac myosin light chain. The hybrid clones that produce specific antibodies were screened by ELISA and RIA methods. The detectable titer of these specific antibodies identified by ELISA and RIA was 1 : 10~4. FACS analysis of 3-D showed that, its DNA content was about the amount of DNA of NS-1 plus those of mouse spleen cell. The 3-D_3, 2-D_9, 1-E_(10) and 2-E_(10) cell lines were stable in secreting monoclonal antibodies in tissue culture for more than 6 months. Two different epitopes on the antigen molecule were measured by RIA with two different monoclonal antibodies. The hybridoma cells produced high titer antibodies in mouse ascites.