Abstract: 【Objective】The study aimed to investigate the protective efficacy of the recombinant antigen of TPO18 (pGEX-TPO18) from Taenia pisiformis. 【Method】 Total RNA was extracted from oncosphere of T. pisiformis. A pair of primers was designed based on the conserved region of other Taeniidae 18 kD gene. The EcoR I and Xho I restriction sites were introduced into 5′ end of primers. After RT-PCR, amplification product was cloned into pMD18-T, sequenced and the sequence analysis was made with bioinformatics. The products were ligated into the pGEX-4T-1 vector after digestion with EcoRⅠ＋XhoⅠ and purified. The recombinant pGEX-TPO18 plasmid was transformed into E. coli BL21 and spots were picked after shaking bacteria. The correctly identified recombinant plasmid, identified by PCR and sequenced, was named pGEX-TPO18. The recombinant plasmids were induced for expression with IPTG, cells were collected by SDS-PAGE analysis and purified using GST agarose resin. Then, TPO18 protein was detected by Western blotting. Rabbits were immunized with the purified recombinant protein emulsified with Freund’s adjuvant, 206 adjuvant, Al(OH)3 adjuvant, respectively, and rabbit anti-TPO18 serum was prepared. Each rabbit was injected with 50 μg recombinant protein for 3 times. On the 34th day after final inoculation, each rabbit was challenged by 1 500 eggs of T. pisiformis. On the 58th day after infection, rabbits were sacrificed and the number of cysts was counted. Before and after 7 days of every immunization, serum was separated. And serum antibody levels were detected using ELISA assay with 40 µg•mL-1 recombinant protein-coated microtiter plates, 1﹕200 dilution of serum and detection of the OD492nm value of serum samples was made.【Result】 The product of RT-PCR was 339 bp and agree with expectations; The results of sequencing showed no mutation. So, the construction of recombinant plasmid pGEX-TPO18 was successful. The pGEX-TPO18 was transformed into E. coli BL21. SDS-PAGE showed that the 38.6 kD fusion protein was expressed at a high level in E. coli. Western blotting analysis showed that a significant response band at 38.6 kD was displayed between rabbit against Cysticercus pisiformis positive serum and recombinant protein. This result indicated that the recombinant protein has reactogenicity and recognized by sera. On the 43th day after final immunization, the levels of antibody from tested group showed a peak. The reduction rate of cysts in immunized sheep (Freund’s adjuvant, 206 adjuvant and Al(OH)3 adjuvant) was 79.13%, 65.66%, and 50.43% respectively. 【Conclusion】Freund’s adjuvant emulsified recombinant antigens showed high protective activity, thus providing valuable information for the development of highly efficient recombinant vaccines against cysticercoids.

Key words: Taenia pisiformis , TPO18 , prokaryotic expression , protective efficacy