金黄色葡萄球菌血浆凝固酶抗血清制备及ELISA检测-生物工程专业论文.docxVIP

Preparation Preparation of Polyclonal Antibodys against StaphyIococcus aureffs and Establishment of ELISA Methods Abstract Staphk7lococcus aureus(S aureus)．belongs to staphylococcus in microbial taxonomy，1S a representative of gram positive bacteria and kind of important human pathogenic microorganism,which lots of serious infections．Therefore，the inspection of S aureus is an important test itern in medicine，health care products，cosmetic and tood· The common test methods ofS．aureus are national standard，fully automatic biochemical method and real-time fluorescence quantificative PCR(RT-qPCR)．Fully automatic biochemical method and RT—qPCR are fast，accurate and realize high-throughput screen． The high price of instruments and consumable materials is limited for basic level inspection organizations．National standard method is the most widely used due to low cost．Rabbit plasma is uesed to make plasma coagulase test and confirm the result，and the stability and controllability ofrabbit plasma iS worse，so there are more false—negative and false positive m Dlasma coagulase test．Therefore，the development of ELISA test method，increased the sensitivity and specificity of plasma coagulase test，could be desired．ELISA test method can also be directly used for rapid screening of S．aureus．The inactivated S．aureus was used as antigen to immunize the N ew Zealand white rabbits，guinea pig．The rabbit antiserum with 1：4000 titer and guinea pig antiserum with 1：2000 titer were obtained．We used the two antiserums and established double-antibody sandwich ELlSA test method，and optimized the experiment condition o f double—antibody sandwich ELISA test method．The optimal experimental conditions were as follow：guinea pig antiserum as coated antibody，rabbit antiserum as detecting antibody，goat anti—rabbit immunoglobulin marked by horseradish catalase as enzyme anti-antibody，rabbit antiserum is diluted at the rate of 1：2000，the rate of guinea pig antiserum is 1：1 000 and the rate of goat anti—r