银屑病自然免疫噬菌体单链抗体库的构建及初步验证  

Construction and Preliminary Validation of a Phage Displayed Human Single-chain Antibody Library Derived from Patients with Psoriasis

作　　者：张银川[1] 张爱华[1] 潘勇兵[1] 余玲[2] 姜功平[2] 方志正[1] 

机构地区：[1]武汉生物制品研究所免疫学研究室,武汉430060 [2]中国人民解放军第195医院皮肤科,湖北咸宁437100

出　　处：《中国生物制品学杂志》2011年第5期502-506,共5页Chinese Journal of Biologicals

摘　　要：目的构建银屑病自然免疫噬菌体展示全人源单链抗体库,并对其进行抗人肿瘤坏死因子α(Humantumornecros-is factorα,hTNFα)单链抗体的富集筛选。方法采用间接ELISA法对33例银屑病患者血清进行抗hTNFαIgG分析;利用噬菌体展示技术,以分离的PBLs构建银屑病患者的pComb3X_scFv抗体库,并对其库容量、正确性和多样性进行鉴定;以rhTNFα包被酶标板,经4轮常规的"吸附-洗脱-扩增"进行特异性hTNFα抗体的固相生物淘选。结果与健康个体相比,银屑病患者具有明显增高的血清抗hTNFα自身抗体(P<0.01);以此抗体基因资源成功构建出多样性良好的单链可变区噬菌体抗体库,库容量约为4×106,重组率>90%;经rhTNFα筛选获得了特异性噬菌体抗体的富集。结论利用构建的银屑病自然免疫噬菌体抗体库可筛选到抗hTNFα特异性单链抗体。Objective To construct a phage displayed human single-chain antibody library derived from patients with psoriasis and screen the specific single-chain antibody against human tumor necrosis factor α（hTNFα）.Methods The serum samples from 33 patients with psoriasis were analyzed for IgG against hTNFα by indirect ELISA.The pComb3X_scFv library was constructed by phage displayed technique using the separated peripheral blood lymphocytes（PBLs）,and identified for capacity,accuracy and diversity.Microtiter plate was coated with hTNFα,based on which the solid phase biopanning of specific hTNFα antibody was performed by 4 cycles of＂adsorption-elution-amplification＂.Results The anti-hTNFα IgG level in sera of patients with psoriasis was significantly higher than that in healthy individuals（P 0.01）.A phage displayed human single-chain antibody library with a capacity of about 4 × 106,a recombination rate of more than 90% and good diversity was successfully constructed using the gene materials of the specific antibody.Specific phage displayed antibody was enriched by biopanning of hTNFα.Conclusion Specific single-chain antibody against hTNFα was successfully screened by using the constructed phage displayed human single-chain antibody library derived from patients with psoriasis.

关 键 词：银屑病 噬菌体抗体库 单链抗体 肿瘤坏死因子Α 

分 类 号：R593.2[医药卫生—内科学] R392-33[医药卫生—临床医学]