关键词: 单增李斯特菌；多克隆抗体;乳胶凝集试验

Abstract:

Abstract:Objective
To prepare the anti-Listeria monocytogenes(LM) polyclonal antibody derived from rabbit and to study the application of LM in latex agglutination test(LAT).Methods
Standard LM strain were cultured and multiplied using selective agents and enrichment  procedures.Freund’s complete inactivated vaccine and Freund’s incomplete inactivated vaccine were made for immunizing rabbits.Blood samples were collected from the marginal ear vein and the antibody titers were measured by ELISA.Four times after immunization,blood serum was separated from the carotid artery and crude IgG was purified  by using the method of caprylic acid-sulfide saturated soution precipitation.IgG molecules were purified by using the HiTrap Protein G chromatography column on AKTA Protein purification device and identified using the methods of SDS-PAGE electrophoresis,UV spectrophotometry,and indirect ELISA.Purified IgG were covalent conjuncted with latex microspheres and the content of latex and interaction time were optimized.The method was established for rapid detection of LM in simulated samples.Results The protein content of the purified anti-LM IgG was 23.364 g·L^-1 and its antibody titer was 1∶12　800-1∶256 00,no cross-reaction occured with a variety of bacteria such as Vibrio cholerae,Salmonella,Shigella,but a weak cross-reaction with Steinmann Listeria monocytogenes.The covalent conjuncted ratio of IgG on the latex microspheres was 1∶40 and the positive rate was 87.5% for detetion in 246 samples by the method of LAT.The minimum detectable content of the antigen was 0.039 8 g·L^-1.Conclusion LAT is a promising method to facilitate the grassroots level for field testing LM with its specificity,high sensitivity and good reproducibility.

Key words: Lislefia monocylogenes;polyclonal antibody;latex agglutination test