副溶血性弧菌鞭毛蛋白FlaA的重组表达与多克隆抗体制备
Recombinant expression and polyclonal antibody preparation of Vibrio parahaemolyticus flagellin A
副溶血性弧菌是广泛存在于水产品中的革兰阴性致病菌。鞭毛蛋白FlaA是副溶血性弧菌所特有的蛋白并在其侵染宿主、引起宿主免疫反应过程中起到重要作用，由鞭毛蛋白免疫动物得到的抗体可以作为检测副溶血性弧菌污染的探针。本研究通过分子生物学方法构建了重组FlaA的表达载体pET28a-FlaA，经IPTG诱导获得FLaA的大量表达。该重组蛋白以可溶性蛋白形式存在，经Ni-NTA亲和柱纯化后对家兔进行免疫，得到浓度为24.32 mg/mL、效价为1:125,000的FlaA抗体。通过间接ELISA实验，测得该抗体与副溶血性弧菌反应的灵敏度为1E4 cfu/mL，且该抗体与五种其它常见致病菌无明显交叉反应。
he Gram-negative bacterium Vibrio parahaemolyticus is a common pathogenic bacteria in aquatic products. Flagellin A, which is a unique protein in this strain, plays an important role during infection and induction of the host immune response. The production of polyclonal antibody against FlaA protein can be used as a probe in V. parahaemolyticus fast detection. The recombinant plasmid pET28a-FlaA was constructed and the FlaA was expressed with induction of IPTG. The soluble FlaA protein was purified by Ni-NTA affinity chromatography and the polyclonal antibody against FlaA was prepared by immunning rabbit. The final concentration of FlaA antibody was 24.32 mg/mL with the titer of 1:125,000. The result of FlaA polyclonal antibody sensibility is 1E4 cfu/mL. The FlaA polyclonal antibody showed no cross-reaction with 5 other kinds of pathogenic bacteria.
夏雨、王燕、王周平
分子生物学微生物学生物工程学
副溶血性弧菌鞭毛蛋白克隆表达抗体制备
Vibrio parahaemolyticusFlagellin Arecombinant expressionpreparation of ployclonal antibody
夏雨,王燕,王周平.副溶血性弧菌鞭毛蛋白FlaA的重组表达与多克隆抗体制备[EB/OL].(2013-03-01)[2025-08-02].http://www.paper.edu.cn/releasepaper/content/201303-36.点此复制
评论