Preparation of mouse polyclonal antibodies against lipoprotein SPD_1609 in Streptococcus pneumoniae

Objective To prepare mouse polyclonal antibodies against lipoprotein SPD_1609 in Streptococcus pneumoniae. Methods Firstly, the spd_1609 gene in Streptococcus pneumoniae (S. pneumoniae) D39 strain was amplified and ligated into the prokaryotic expression vector of pGEX-4T-1 to generate recombinant plasmid pGEX-4T-1609. Secondly, the fusion protein of GST-1609 was expressed in Escherichia coli BL21 (DE3), and purified using GST affinity chromatography. After cleaving the GST-tag of the fusion protein, the SPD_1609 protein was obtained and purified using GST affinity chromatography. Finally, BALB/c mice were immunized with the purified SPD_1609 protein to produce polyclonal antibodies. The titer and specificity of polyclonal antibodies were detected using ELISA and Western blotting. Results In this study, the recombinant vector of pGEX-4T-1609 was constructed, and the SPD_1609 protein (Mr is approximately 35 000) was expressed and purified with a higher 95% purity. The ELISA results suggested that the SPD_1609 protein could induce specific immune responses in mice, and the titer of antibody in the immunized mice was 1:40 960. Western blotting indicated that the polyclonal antibody could specifically recognize the SPD_1609 protein expressed in prokaryotic expression vector and in S. pneumoniae. Conclusion The mouse polyclonal antibody with good specificity against SPD_1609 protein has been successfully obtained.