目的使用构建轻链次级库的方法,对已经获得的人源抗TNFα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TNFαmAb的Fab。方法首先,采用RTPCR扩增正常人全套抗体轻链基因,并与已获得的人源抗TNFαmAb的重链基因配对,构建人源抗TNFα噬菌体抗体的轻链次级库。然后筛选与TNFα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TNFαmAbFab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。 OBJECTIVE: To construct a light chain secondary library, a light chain replacement was performed on the Fabs of the human anti-TNF mAb mAbs that have been obtained, and the Fabs with higher affinity human anti-TNF-α mAbs were screened. Methods Firstly, RT-PCR was used to amplify a full set of normal human light chain gene and paired with the heavy chain gene of human anti-TNFα mAb to construct a light chain secondary library of human anti-TNFα phage antibody. Then screening and TNF  α has a higher affinity of the clone. Results After three rounds of bio-panning biopanning, human antibody Fab with higher affinity than the original human anti-TNF-α mAb Fab was obtained. Conclusion The light chain secondary library screening method can effectively improve the affinity of the antibody, providing an effective method to solve the problem of low affinity of the human antibody prepared by the phage antibody library method.