摘要
通过轻链替换，提高抗ＨＢＳＡｄｅ菌体抗体的结合力。方法将ＰＣＲ扩增的人抗体轻链基因，插至已克隆有人源抗乙肝表面抗原（ＨＢｓＡｇ）抗体重链Ｆｄ基因的噬菌粒中，构建成轻链替换文库，通过亲和淘洗，筛选与Ｆｄ相匹配的轻链基因。结果经亲和淘洗后，筛选出了与Ｆｄ相匹配的轻链基因，链替换后，噬菌体抗体（ＰｈＡｂ）的ＥＬＩＳＡ光吸收值（Ａ）从０．４３±０．０９提高到最高为１．２４±０．１０。抑制实验证实所筛选出来的ｎｈａｂ具有抗ＨＢｓＡｚ的特异性。分析了ＥＬＩＳＡ光吸收值（Ａ）最高的５株噬菌体抗体的轻链基因序列，结果表明，３株ｋ链的碱基序列一致，属ＶｋⅢ亚群；２株链的基因序列相同，均属ＶＩ亚群。结论结果表明经链替换的噬菌体抗体结合力得到了较大提高，并具有与ＨＢｓＡｇ合的特异性。
Objective We had cloned an Fd gene of human antibody against HBsAg previously. Theobjective of this experiment is to improve the affinity phage antibody by chain-shuffling. MethodsHuman antibody light chain gene repertoire was generated by RT-PCR from human peripheral bloodlymphocyte, and then phage antibody sublibrary was constructed by inserting the repertoire into thephagmid which contained the Fd gene. The matched gene of light chain was selected by biopaning.Results After three rounds of selection, eight clones with higher absorbance than that of original cloneat 490 urn in ELISA were obtained. DNA sequencing showed three of the five W genes were x type, andthe other two were X type. Conclusion The results indicated that the affinity of chain shuffled phageantibodies(phab) were improved. The specificity of the selected phab was also confirmed.
出处
《中华肝脏病杂志》
CAS
CSCD
1999年第1期36-38,共3页
Chinese Journal of Hepatology
基金
国家863计划重大项目资助!Z18-01
