Abstract: Abstract：Objective To verify the role of nanobodies in epitope display by displaying the C-terminal epitope of β-HCG in the CDR3 region of nanobodies. Methods The C-terminal epitope of β-HCG (amino sequence 151-165) was inserted into the CDR3 region of the nanobody by gene synthesis method. The prokaryotic expression vector pET24a was constructed by digestion and ligation, and the expression was induced by IPTG. His tag filler was purified. The purified target protein was immunized to New Zealand white rabbits. After five immunizations, the indirect ELISA titer was detected and the polyclonal antibody was purified by antigen-coupled purification medium. Western blot assay was used to detect the specificity of polyclonal antibodies. Results Prokaryotic expression vector was constructed, and the highly expressed strains were obtained. After induction by IPTG, the target protein existed mainly in the form of inclusion bodies. Affinity chromatography obtained the target protein with purity greater than 98%. After refolding, the inclusion bodies were immunized with titer of 1∶512 000. Western blot specific detection showed that the immune polyclonal antibody could specifically bind to β-HCG. Conclusion The method of displaying the C-terminal epitope of β-HCG antigen in CDR3 region of nanobody can be used for the preparation of β-HCG antibody, and which lay a foundation for the future research of displaying the epitope of nanobody.

Key words: antibodies, chorionic gonadotropin, epitopes, nanobody, antibody preparation