融合基因Aβ-HBcAg的原核表达及表达蛋白的免疫反应性和免疫原性分析

目的　研究 β 淀粉样肽 (β amyloidpeptide,Aβ)与乙型肝炎核心抗原 (HBcAg)融合基因Aβ HBcAg的原核表达 ,检测融合蛋白的免疫反应性及免疫原性。方法　将重组质粒pBV2 2 0 /Aβ HBcAg转化大肠杆菌DH5α,温控表达 ,超声破碎细菌 ,通过SDS PAGE、考马斯亮蓝染色和ELISA等方法 ,观察Aβ HBcAg的表达及融合蛋白的免疫反应性 ;用融合蛋白免疫Balb/c小鼠 ,ELISA法检测血清中的抗 Aβ抗体和抗 HBc抗体滴度。结果　融合蛋白以包涵体的形式存在于沉淀中 ,表达量为菌体沉淀的 5 % ,融合蛋白既有Aβ的免疫反应性 ,又有HBcAg的免疫反应性。Balb/c小鼠经过 3次免疫后 ,血清中的抗体滴度很低 ,继续强化免疫 2次后 ,血清中抗 Aβ抗体滴度、抗 HBc抗体滴度分别上升为 1∶80 0和1∶32 0 0。结论　融合基因Aβ HBcAg可在大肠杆菌DH5α中表达 ,表达蛋白有一定的免疫反应性和免疫原性 ,但表达量较低 ,免疫反应性和免疫原性较弱。提示融合基因Aβ HBcAg的表达量和融合蛋白免疫原性的提高尚需进一步研究。

Prokaryotic expression of fusion gene Aβ-HBcAg and analysis of the immunoreactivity and immunogenicity of the expression protein

Objective To study the prokaryotic expression of fusion gene Aβ-HBcAg and analyze the immunoreactivity and immunogenicity of expression protein. Methods Recombinant plasmid pBV220/Aβ-HBcAg was transformed into E.coli DH5α, and expressed by temperature inducing. The bacteria were split by ultrasonic wave. The expression of the fusion protein was studied by SDS-PAGE and Coomassie brilliant blue staining. The immunoreactivity of the fusion protein was determined using ELISA. After immunized intraperitoneally with the fusion protein, 5 Balb/c mice's sera titers of anti- Aβ and anti-HBc were evaluated by ELISA. Results Fusion protein was in sediment of the split bacteria as inclusion bodies and its expression level was 5% of the total sediment protein. The fusion protein had both immunoreactivity of Aβ and HBcAg. The titers of anti-Aβ and anti-HBc were very low after 3 times of immunization. After immunization for 5 times, the titers reached 1∶800 and 1∶3 200 for anti-Aβ and anti-HBc, respectively. Conclusion Recombinant gene Aβ-HBcAg can be expressed in E.coli DH5α and the expression protein has certain immunoreactivity and immunogenicity. It indicates that further work should be done to enhance the expression level of fusion gene Aβ-HBcAg and improve the immunogenicity of the fusion protein.