刀豆凝集素单克隆抗体特性分析和初步应用

建立了一株能分泌抗刀豆凝集素(ConA)单克隆抗体(McAb)的细胞株-CA2。ELISA证实所得 CA2 McAb仅与 ConA有反应,而与菜豆凝集素(PHA)、美洲商陆(PWM)、花生凝集素(PNA)、扁豆凝集素(LcA)以及牛血清白蛋白(BSA)都无交叉反应。免疫印迹试验进一步证实该McAb 只能与 ConA 形成免疫沉淀线。该McAb 能抑制 ConA对RLc-802细胞的凝集作用,竞争抑制试验显示α-甲基甘露糖、果糖不能抑制 CA2 McAb与 ConA 的结合,提示它可能是针对 ConA蛋白结合点部位抗原决定簇的 McAb。应用该McAb证实了急性T淋巴细胞白血病细胞株Molt_4、CEM和B淋巴母细胞CCRF-sb以及转化的扁桃腺淋巴细胞表面膜存在有分子量大小不同的 ConA受体,提示同是ConA受体,可能具有不同的生物学功能。

CHARACTERIZATION OF MONOCLONAL ANTIBODY AGAINST CONCANAVALIN A AND ITS PRELIMINARY APPLICATION

A new cell line CA2, secreting a monoclonal mouse IgG1, an antibody specific for protein concanavalin A (Con A),has been established. The apparent molecular weights of heavy and light chains from purified antibody determined by NaDodSO4/PAGE was 56,000 and 27,000 daltons respectively. CA2 was screened and characterized by BLISA using a variety of lectins. Among the various lectins,CA2 reacted only with Con A, and no cross reactivity was observed with PHA,PWM,PNA,LcA and BSA as well.Inhibition of agglutination test and competitive inhibition assay studies showed that CA2 can inhibit the agglomerative effect of Con A on rat hepatocellular carcinoma cell line KLC-802 and competitive inhibition of binding of CA2 was not achieved withα-methylmannoside and fructose, suggested that the specificity of CA2 monoclonal antibody may be directed against the protein binding sites for Con A.By using CA2 monoclonal antibody as an analytical probe for the Con A receptors on the cell membrane and at the same time it was also explored by SDS-PAGE, and Immunobiotting method. It was found that Con A receptors of various molecular weight do existed on the cell membranes of acute T-cell lymphocytic leukemia cell lines (Molt4), CEM and B-lymphoblastic cell line (CCEF-Sb)as well as on transformed tonsillar lymphocytes. It is possible that Con A receptors because of their difference in molecular weight may express different biological functions.