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《Xinjiang Agricultural Sciences》 2017-07
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Rapid Generation of Anti-FSHR Nanobody by CDR3 Affinity Transfer Approach

XI Ou-yan;QIU Ling-ling;MA Xiao-ling;QIN Rui-ping;ZHAO Ting;LI Jiang-wei;Xinjiang Key Laboratory of Biological Resources and Genetic Engineering/College of Life Science and Technology,Xinjiang University;  
【Objective】To evaluate the potential of camel-derived antibody c Ab BCII10 as a non-antibody affinity transfer skeleton,the FSHR binding peptide was grafted to the antigen-binding region of c Ab BCII10 using affinity transfer to rapidly obtain anti-FSHR antibody.【Method】The FSH binding motif FSH 33-53 coding sequence was inserted into the CDR1 and CDR3 regions of the nanobody c Ab BCII10,respectively,and named VHH-h FSH1 and VHH-h FSH3.These DNA sequences were cloned into p ET22 b vector and transformed into E.coli BL21( DE3),and purified single-domain antibody was obtained by IPTG induction and Ni-affinity affinity chromatography.【Result】Finally binding capacity and specificity of purified c Ab BCII10,VHH-h FSH1 and VHH-h FSH3 with FSHR were identified by ELISA.The VHH-h FSH1,VHH-h FSH3 and c Ab BCII10 proteins obtained by the framework transplantation were expressed in the intercellular space of bacteria.ELISA experiments showed that the VHH-h FSH3 obtained by grafting the FSHR binding peptide FSH33-53 to the CDR3 of c Ab BCII10 had a specific binding to FSHR activity.【Conclusion】CAb BCII10 can be used as a graft framework,FSH and FSHR binding peptide grafted into the CDR1 and CDR3 regions of c Ab BCII10 can obtain higher affinity anti-FSHR antibody.
【Fund】: 国家自然科学基金项目“骆驼单域抗体框架区作为小分子抗体通用移植骨架的研究”(31370933);国家自然科学基金项目“基于高通量测序对骆驼抗体库的多样性分析和数据挖掘及新纳米抗体的发现”(31570935)~~
【CateGory Index】: R392
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