摘要
目的 :以MUC1为靶点研制抗肿瘤蛋白疫苗。方法 :将MUC1基因连入pMAL p2原核表达载体 ,并转化大肠杆菌 ,通过IPTG诱导MUC1表达 ,经Westernblot鉴定 ,Amylose亲和层析纯化蛋白。用MUC1 MBP免疫健康C5 7BL 6小鼠 ,测定其免疫活性 ,通过ELISA测定血清中抗MUC1抗体的效价 ;采用MTT法测定小鼠脾CTL活性 ,通过3H TdR掺入法测定T细胞增殖能力。结果 :成功构建了pMAL MUC1表达载体并得到稳定表达MUC1的菌株 ,鉴定了MUC1在大肠杆菌中的表达 ,纯化了MUC1蛋白。经重组MUC1 MBP免疫的C5 7小鼠 ,血清抗MUC1抗体的效价为 1:5 76 0± 32 2 1;脾CTL对MCF 7及Lewis肺癌细胞的杀伤率分别为 4 7 7%± 4 3%和 6 7 5 %± 6 5 %。结论 :人类重组MUC1融合蛋白可引发小鼠CTL反应和体液免疫应答 ,有希望研制成抗腺癌蛋白疫苗。
Objective:To study MUC1 based cancer vaccine.Methods:MUC1 gene was inserted into pMAL-p2 vector and constructed recombinant pMAL-MUC1. MUC1-MBP fusion protein expression was induced by IPTG in E coli DH5α transformed by the recombinant pMAL-MUC1 .The fusion protein was analyzed by Western blot and purified by amylose affinity chromatography. The antiserum,T cell proliferation and CTL activity of spleen from C57 mice immunized by MUC1-MBP were determined respectively by ELISA,adding 3H-TdR and MTT.Results:Had successfully constructed pMAL-MUC1 expression vector,and purified MUC1-MBP and MBP. C57 mice immunized by MUC-MBP generated MUC1 specific antibody and CTL.The titer of polyclonal antibody to MUC1 was about 1∶5 760±3 221. CTL cytotoxicity to the MCF7 and lewis lung cancer cells respectively were at 47.7%±4.3% and 67.5%±6.5%.Conclusion:Human recombinant MUC1-MBP fusion protein activated T and B cell response in mice.The results suggested that the recombinant Muc1 may be used to develop protein vaccine against carcinoma.
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2003年第11期747-751,共5页
Chinese Journal of Immunology
基金
国家自然科学基金项目 (NO3 9670 686)资助
