自制多抗检测金黄色葡萄球菌PVL蛋白的研究

Study of PVL Protein Detection in Staphylococcus aureus by Self-Made Polyclonal Antibody

摘要: 目的：携带杀伤白细胞素(pvl)基因的耐甲氧西林的金黄色葡萄球菌毒力强，可以诱发高致死率的社区获得性肺炎，但目前仍缺少快速简便的检测方法。本研究拟制备抗PVL多克隆抗体用于其快速检测。方法：本研究通过将金黄色葡萄球菌PVL蛋白S亚基的重组质粒LukS-PV/pET28a导入大肠杆菌诱导表达重组蛋白，再经免疫新西兰兔制备抗血清，以Western blotting法检测金黄色葡萄球菌PVL蛋白表达。结果：成功纯化出重组LukS-PV蛋白，并制备抗LukS-PV抗血清。通过Western blotting法采用自制多克隆抗体成功检测出金黄色葡萄球菌PVL表达。结论：自制抗PVL多克隆抗体可以检测金黄色葡萄球菌PVL表达，为下一步试剂盒制备奠定技术基础。

Abstract: Objective: Methicillin-resistant Staphylococcus aureus (MRSA), which carries the Panton-Valentine leukocidin (pvl) gene, is highly virulent and can cause community-acquired pneumonia with a high fatality rate, but a quick and easy test is still lacking. In this study, polyclonal antibodies against PVL were prepared for rapid detection. Methods: In this study, the recombinant plasmid LukS-PV/pET28a of PVL protein S subunit of S. aureus was introduced into Escherichia coli to induce the expression of the recombinant protein, and then anti-serum was prepared by immunizing rabbits, and the expression of PVL protein of S. aureus was detected by Western blotting method. Results: Recombinant LukS-PV protein was successfully purified and antiserum was prepared. The expression of PVL in S. aureus was detected by Western blotting and polyclonal antibody. Conclusion: The self-made anti-PVL polyclonal antibody can detect the expression of PVL in S. aureus, and lay a technical foundation for the preparation of the kit.