Abstract: Objective To explore the immune effect of the outer membrane protein A ( OmpA ) prokaryotic expression protein of Rodentibacter heylii ( Rh ) on mice. Method Prokaryotic expression and purification by nickel ion affinity chromatography were used to prepare Rh OmpA recombinant protein, and the target protein was identified by SDS-PAGE and Western blot. The obtained OmpA target protein was immunized to ICR mice by intraperitoneal and intramuscular injection and nasal drip. Subsequently, the antibody titer was determined by ELISA method . Then Rh ATCC12555, Rodentibacter pneumotropicus (Rp) ATCC35149, and Rh isolate PP320 were challenged to the immunized mice abdominal cavity to evaluate the protective effect of recombinant protein OmpA on mice. Result The OmpA specific antibody level in serum of immunized mice was determined by ELISA with the expression protein OmpA as the coated antigen. The average antibody titer was 19. 84log₂. The protective rates of immunized mice against two standard strains and PP320 were 60%, 60% and 80%, respectively. The mortality rate was significantly reduced(P<0. 001) . The ELISA method based on OmpA expressed protein detects the sera of Pasteurella pneumotropica( Pp) positive mice, and the antibody positive rate is 60%. Conclusion The prepared Rh OmpA expressed protein had good immunogenicity and had a certain protective effect on Rh and Rp, which can provide a reference for the prevention and control of respiratory pathogens in laboratory animals.

Key words: Rodentibacter heylii, OmpA, prokaryotic expression, evaluation of immune efficacy

摘要： 目的 探讨海立啮齿杆菌( Rh) 外膜蛋白 A( OmpA) 原核表达蛋白对小鼠的免疫效果。 方法 采用原核表达、镍离子亲和层析纯化,制备 Rh OmpA 重组蛋白,并用 SDS-PAGE 和 Western blot 进行鉴定。 获得的 OmpA 目的蛋白经腹腔、肌肉联合注射和滴鼻两种方案免疫ICR小鼠, 由 ELISA方法测定其抗体滴度。再分别将 RhATCC12555、嗜肺啮齿杆菌(Rp)ATCC35149 和 Rh 分离株 PP320 对免疫小鼠腹腔接种攻毒,评价重组蛋白 OmpA对小鼠的保护效果。 结果 以表达蛋白 OmpA 为包被抗原的 ELISA 方法检测免疫小鼠血清中OmpA 特异性抗体水平,平均抗体滴度为 19. 84log2。 免疫小鼠对两标准菌株和 PP320 的保护率分别为 60%、60%和 80%,与未免疫小鼠相比死亡率显著降低(P<0. 001) 。 基于 OmpA 表达蛋白建立的 ELISA 方法检测嗜肺巴斯德杆菌( Pp) 阳性小鼠血清,抗体阳性率为 60%。 结论 制备的 Rh OmpA 表达蛋白有良好的免疫原性,对 Rh 和 Rp 具有一定的保护效果,为实验动物中呼吸道病原菌的防控提供参考。

关键词: 海立啮齿杆菌, 外膜蛋白 A, 原核表达, 免疫效果评价