含1型血小板结合蛋白基序的去整合素样金属蛋白酶2(ADAMTS2)多克隆抗体的制备  

Preparation of rabbit polyclonal antibody against mouse ADAMTS2

作　　者：付俊[1] 缪时英[1] 王琳芳[1] 

机构地区：[1]中国医学科学院基础医学研究所,北京协和医学院基础学院,医学分子生物学国家重点实验室,生物化学与分子生物学系,北京100005

出　　处：《细胞与分子免疫学杂志》2016年第3期393-396,401,共5页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(31401273);国家重大科学研究计划(2012CB944902)

摘　　要：目的原核表达并纯化小鼠源性含1型血小板结合蛋白基序的去整合素样金属蛋白酶2(ADAMTS2)蛋白的C段(1109-1213),制备兔抗ADAMTS2多克隆抗体。方法以重组质粒p GEX-6p-1-ADAMTS2(1109-1213)转化大肠杆菌,异丙基-β-D-硫代半乳糖苷(IPTG)诱导目的蛋白表达,通过亲和纯化,并且经质谱鉴定;以表达的目的蛋白免疫新西兰大白兔,制备抗ADAMTS2的多克隆抗体。ELISA检测抗体效价,Western blot法鉴定抗体特异性。结果在大肠杆菌中表达出目的蛋白ADAMTS2,纯化后经质谱鉴定并免疫新西兰大白兔,成功获得抗血清。血清效价达到1∶160 000以上,且具有良好的特异性。结论成功制备出效价高、特异性好的兔抗ADAMTS2抗体。Objective To obtain recombinant mouse col agenase ADAMTS2（ ADAM metal opeptidase with thrombospondin type 1 motif,2） C terminal（ 1109-1213）,and prepare the corresponding rabbit anti-ADAMTS2 polyclonal antibodies. Methods The recombinant expression plasmid p GEX-6p-1-ADAMTS2（ 1109-1213） was transformed into E. coli. The target protein was induced by IPTG and identified by mass spectrometry following affinity purification. The expressed and purified ADAMTS2（ 1109-1213） protein was used to immunize New Zealand rabbits to prepare anti-ADAMTS2 polyclonal antibodies. The antibody titers were detected by ELISA and the antibody specificity by Western blotting. Results The protein ADAMTS2（ 1109-1213） was expressed in E. coli after IPTG induction,and with the purified protein,we prepared antiserum in the immunized rabbits. The titer of the antiserum reached over 1∶160 000. The antiserum showed a good specificity. Conclusion The high titer and specific rabbit anti-ADAMTS2 antibody has been prepared successfully.

关 键 词：ADAMTS2 原核表达 多克隆抗体 

分 类 号：R392.11[医药卫生—免疫学] Q786[医药卫生—基础医学]