抗癌胚抗原单区抗体的质粒构建及表达

Research on Anti-carcinoembryonic Antigen Single Domain Antibody of Plasmid Construction and Expression

目的对抗癌胚抗原单区抗体进行表达研究。方法采用PCR技术将已获得的抗癌胚抗原（CEA）鼠源单克隆抗体E7B10重、轻链可变区基因（VH、VK）进行改造后分别克隆进大肠杆菌表达质粒pET－24α（＋）中，并进行表达。结果显示重、轻链可变区在大肠杆菌中获得了高效表达，其表达量达到菌体总蛋白的20％与25％，SDS－PAGE和Westernblot图谱显示表达产物分子量为13KD、12KD，与其基因编码蛋白的理论推算值相符。表达产物在胞内主要以不溶性包含体存在。结论经变性和复性处理后，RIA表明重链可变区基因表达产物具有结合其特异性抗原CEA的能力。

Objective To probe into expression of single domain antibody for anticarcinoembryonic antigen (CEA). Methods The heavy and light chain variable region (VH and VK) genes encoding the murine monoclonal antibody E7B10 with specificity for CEA were amplified by PCR and cloned into expression vector PET-24a(+) by gene recombinant. Results The constructs were expressed in E.Coli with high level as inclusion bodies(IB), reaching 20% and 25% of the total bacteria protein, and characterized by SDS-PAGE and Western blot to have molecula weights of 13KD and 12KD. Conclusion The expression product of the heavy chain variable region has CEA - binding ability in RIA by means of IB solubilization then refolding