摘要
目的使用构建轻链次级库的方法,对已经获得的人源抗TNFα单抗mAb的Fab进行轻链置换,并筛选具有更高亲和力的人源抗TNFαmAb的Fab。方法首先,采用RTPCR扩增正常人全套抗体轻链基因,并与已获得的人源抗TNFαmAb的重链基因配对,构建人源抗TNFα噬菌体抗体的轻链次级库。然后筛选与TNFα具有更高亲和力的克隆。结果经过3轮的生物淘筛biopanning,获得了较原来的人源抗TNFαmAbFab具有更高亲和力的人源抗体Fab。结论轻链次级库筛选法,能有效地提高抗体的亲和力,为解决噬菌体抗体库法制备的人源抗体亲和力较低这一难题,提供了一种有效的方法。
AimToobtainhumananti TNF αantibodyFabswithhigheraffinitythroughreplacingthelightchainofthehumananti TNF αantibodyFabfragmentbyconstructingalightchainsecondlibrary.MethodsFirstlyamplifytherepertoireofthehumanlightchainsgenesfromhumanperipheralbloodlympho-cytesusingRT PCRandassemblethemrandomlywiththeheavychaingeneoftheanti TNFαantibodysoastoconstructasecondlightchainlibrarywhichcomprisesasingleheavychaingeneoftheantibodytoTNFαandallofthehumanlightchaingenes.Thenselecttheantibodycloneswithhigherbindingaffinity.ResultsSomenewphagedisplayedantibodieswhichhadhigheraffinityforTNFαwereobtainedfromthenewlibrarythroughthreeroundsofbiopanning.Conclusionconstructinglightchainsecondlibrarycouldeffectivelyimprovetheaffinityoftheantibodyandcouldbeavalidmethodtoovercomethedis-advantageoftheloweraffinityofthehumanantibodyfromphagedisplay.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2001年第2期179-181,共3页
Chinese Journal of Cellular and Molecular Immunology
基金
国家自然科学基金资助
No.39670043