Abstract: [Objective] To explore the optimal preparation method of recombinant Brucella outer membrane protein Omp19, to evaluate its immunogenicity in experimental animals, and to lay the foundation for the follow-up pilot-scale study. [Method] The recombinant Omp19 protein was expressed by prokaryotic induction using the shake flask culture method, and the effects of different culture mediums, IPTG concentrations, induction temperatures, induction durations, and bacterial densities on protein expression were evaluated and optimized. The recombinant Omp19 protein was expressed and purified by cation exchange chromatography, hydrophobic chromatography, and anion exchange chromatography methods. The purity of the recombinant Omp19 protein was characterized using SDS-PAGE and SEC-HPLC methods. The immunogenicity of the recombinant Omp19 protein was evaluated by animal experiment. [Result] Shake flask culture test showed that the better expression conditions of recombinant Brucella Omp19 protein were as followed: using the LB medium culture and inducing at OD_{600 nm} of 0.6 ~ 1.0 of bacterial densities with 0.5 mmol/L IPTG at 28 ℃ for 5 h. The highly pure recombinant Omp19 protein was obtained after the three steps of purification. Immunogenicity studies revealed that the recombinant Omp19 protein could effectively stimulate the production of specific antibodies when formulated with adjuvants in BALB/c mice. [Conclusion] This research optimized the preparation method of recombinant Brucella Omp19, evaluated its immunogenicity in mice, and provided the experimental basis for recombinant Brucella vaccine development.

Key words: Brucella, outer membrane protein Omp19, preparation method, protein purification, immunogenicity