Abstract: This work aims to construct the prokaryotic protein of glypican3（GPC3）and to prepare its anti-serum to mouse. The complete open reading frame（ORF）of human GPC3’s cDNA was amplified by PCR,and then cloned into prokaryotic expression vector of pET28a. Further,the GPC3 fusion protein was expressed in Escherichia coli BL21,the fusion his-protein was purified by affinity chromatography and the fusion protein of purity was measured by SDS-PAGE electrophoresis. Finally,the protein content was also measured. The Balb/c mice were immunized by the purified protein for preparing the anti-serum,and the anti-serum was identified with ELISA analysis. Results showed that the polyclonal antibody against GPC3 protein was successfully prepared and the titer of antisera was 1：51200;moreover,the specificity was validated to be fine by Western blot. Conclusively,recombinant human GPC3 protein expressed in the prokaryotic system has strong immunogenicity.

Key words: glypican3, fusion protein, polyclonal antibody