重组大肠杆菌不耐热肠毒素B亚单位表达条件的优化及其佐剂作用

Optimization of Condition for Expression of Recombinant E.coli Heat-labile Enterotoxin B Subunit and Adjuvanticity of Expressed Product

目的优化重组大肠杆菌不耐热肠毒素B亚单位(LTB)的表达条件,并检测其黏膜佐剂作用。方法通过向LB培养基中添加葡萄糖(终浓度为0.5%)、低温培养诱导等方法诱导LTB的表达;采用阳离子交换法对目的蛋白进行纯化;以LTB作为佐剂进行免疫试验。结果可溶性重组大肠杆菌不耐热肠毒素(LTB)的表达量明显提高,纯化后蛋白含量达95%以上,经纯化的LTB作为三价流感裂解疫苗或gD抗原的佐剂免疫BALB/c小鼠,可提高小鼠血清及黏膜的抗体水平,且其效果与铝化剂及福氏佐剂相当或更高。结论优化了LTB的表达条件,且表达的LTB具有较强的黏膜免疫佐剂作用。

Objective To optimize the condition for expression of recombinant E. coli heat-labile enterotoxin B subunit （LTB） and determine the adjuvanticity of expressed product.Methods Optimize the condition for expression of rercombinantE, coli LTB by addition of glucose to LB medium to a final concentration of 0.5 % and induction at low temperature. Purify the expressed LTB by cation exchange chromatography. Immunize BALB/c mice with split influenza virus vaccine using LTB as adjuvant and determine the antibody level induced. Results After the condition for expression was optimized, the expression level of LTB increased significantly. The protein content of LTB after purification reached more than 95% .The serum and mucosal antibody levels of mice immunized with split influenza virus vaccine containing LTB as adjuvant were equivalent to even higher those containing aluminum or Freund adjuvant. Conclusion The condition for expression of recombi- nant E. coli LTB was successfully optimized, and the expressed LTB showed good adjuvanticity to mucosal immunization.