摘要
目的对通过大容量抗体库中筛选到的4株抗TNF-人单链抗体进行分离纯化及鉴定。方法采用硫氰酸盐洗脱ELISA法评估阳性克隆的亲和力;亲和层析柱纯化抗体;通过ELISA检测抗体的特异性结合活性;用L929细胞检测其体外中和TNF-α的活性。结果用硫氰酸盐洗脱法测定4个噬菌体抗体的相对亲和力与其相应的可溶性抗体相对亲和力相一致;SDS-PAGE电泳结果表明,单链抗体得到成功纯化;ELISA结果表明,纯化的抗体能够特异性结合人TNF-α;体外中和试验证明筛选到的单链抗体能中和TNF-α对L929细胞的毒性。结论从人源大容量噬菌体抗体库中能够筛选到具有中和活性的抗TNF-α人单链抗体。
Objective To purify, and characterize human anti-TNF-α ScFv obtained from large phage antibody library. Methods The relative affinity of the 4 clones were measured by ELISA using thiocyanate elution. The ScFv has been purified by affinity chromatography. The iu vitro neutralization assay was used to test the biological activity of ScFv. Results The relative affinity of 4 phage antibodies measured by ELISA follow using thiocyanate elution e, oincided with that of the corresponded ScFv. The analysis of SDS-PAGE showed that the ScFv was purified successfully. Functional analysis demonstrated that ScFv could neutralize the cytotoxicity of TNF-α. Conclusion The neutralizing human anti-TNF-αScFv could be obtained frum large phage antihody library.
出处
《免疫学杂志》
CAS
CSCD
北大核心
2008年第6期700-702,共3页
Immunological Journal
基金
北京市自然科学基金(5052025)
军队医药卫生一般专项(06Z007)资助项目
