Preparation and characterization of monoclonal antibody against hTSC29 pep-tides / 中国免疫学杂志

ABSTRACT

Objective:To prepare monoclonal antibody ( mAb ) against human testis-specific conserved gene ( hTSC29 ) peptides and characterize its immunological and biological features.Methods:According to bioinformatics analysis and prediction of the antigenicity, surface property, hydrophilicity and flexibility of hTSC29, a 18-amino acid residue partial peptide of hTSC29 was synthesized,then immunized the BALB/c mice for preparing antiserum.The mAb against hTSC29 was produced using the routine hybridoma technique.The properties of the mAb against hTSC29 were identified by ELISA, Western blot and immunohistochemistry staining.Results:After cell fusion and subcloning, one hybridoma cell lines secreting specific mAb against hTSC29 protein were obtaind.The Ig subclass of the mAb was IgG2b(κ).ELISA detection showed that the titer of mAbs in cultured was 1∶104.Western blot analysis proved that the mAb could specifically recognize Mr 60 000 protein in human testis total protein.The hTSC29 protein main located at circumference of spermatocyte and spermatid in human testis tissue by immunohistochemistry staining and immunofluorescence assay.Conclusion:One hybridoma cell lines which can secrete specific mAb against hTSC29 protein with high titers and specificity have been established successfully.The mAb will provide efficient tools for functional studies of hTSC29 expressed in spermatogenesis.