Monoclonal antibodies against Listeria monocytogenes
YIN Yin1, 2, LIU Su3, WANG Fang2, ZHU Xue-liang2, ZHOU Bang-xin2, LUO Xue-nong2
1.Tianjin Center For Animal Disease Control and Prevention, Tianjin 300402, China; 2.State Key Laboratory of Veterinary Etiological Biology,Lanzhou Veterinary Research Institute, CAAS,Lanzhou 730046, China; 3.Lanzhou Institute of Biological Products, Lanzhou 730046, China
Abstract：The aim of this study was to prepare monoclonal antibodies against P60 and develop a double-antibody sandwich enzyme-linked immunosorbent assay, which would be used in the detection of Listeria monocytogenes. The purified recombinant protein of P60, produced by prokaryotic expression, was used to immunize the Balb/c mice. The hybridoma cell lines against P60 were obtained by fusing spleen cells from the immunized mice with mouse Sp2/0 myeloma cells. Specific McAb against L. monocytogenes was screened by ELISA. The isotype of the McAb was identified, and then the titers, stability and the affinity of ascites were analyzed. Sandwich ELISA was established using purified ascites and biotinylated P60 polyclonal antibody to detect the p60, which obtained from supernatant of Listeria cultures. The standard curve was established for quantitative determination using the recombinant P60 as the standard against the corresponding the value of OD450. Results showed that 6C2 McAb could be recognized specifically by L. monocytogenes, and the ELISA method based on 6C2 McAb could be used in identification and quantitative detection of samples contaminated with L. monocytogenes.
殷茵, 刘溯, 王芳, 朱学亮, 周邦信, 骆学农. 单增李斯特菌P60单克隆抗体的制备及初步应用[J]. 中国人兽共患病学报, 2015, 31(6): 527-531.
YIN Yin, LIU Su, WANG Fang, ZHU Xue-liang, ZHOU Bang-xin, LUO Xue-nong. Monoclonal antibodies against Listeria monocytogenes. Chinese Journal of Zoonoses, 2015, 31(6): 527-531.
[1] Dussurget O, Pizarro-cerda J, Cossart P. Molecular determinants of Listeria monocytogenes virulence[J]. Annu Rev Microbiol, 2004, 58(6): 587-610. doi:10.1146/annurev.micro.57.030502.090934 [2] Bielecki J. Emerging food pathogens and bacterial toxins[J]. Acta Microbiol Pol, 2003, 52(Suppl): 17-22. [3] Becker B, Jorda S, Holzapf WH, et al. Rapid and specific detection of Listeria monocytogenes in smoked salmon with BAX-PCR[J]. Food Ctrl, 2005, 16(8): 717-721. [4] Xu DS, Shen YH, Yu MH. Development of a rapid,sensitive and specific polymerase chain reaction method for detection of Listeria monocytogenes in foods[J]. Mod Prevent Med, 2007, 34(16): 3040-3041. (in Chinese) 徐德顺, 沈月华, 俞明华. 食品中单核细胞增生李斯特菌PCR快速检测方法[J]. 现代预防医学, 2007, 34(16): 3040-3041. [5] Boerlin P, Boerlin PF, Jemmi T. Use of listeriolysin O and internalin A in a seroepidemiological study of Listeriosis in Swiss dairy cows[J]. J Clin Microbiol, 2003, 41(3) : 1055-1061.doi:10.1128/JCM.41.3.1055-1061.2003 [6] Vazquez-boland JA, Dominguez-bernal G, Gonzalez-zorn B, et al. Pathogenicity islands and virulence evolution in Listeria [J]. Microbes Infect, 2001, 3: 571- 584. [7] Cui HZ, Zhang H, Wand XL. Preparation of monoclonal antibody to Listeria monocytogenes and development of colloidal gold strip for detection of the bacterium[J]. Chin Vet Sci, 2010, 40(01): 45-50. (in Chinese) 崔焕忠, 张辉, 王兴龙. 单增李斯特菌单克隆抗体的制备及胶体金试纸的研制[J]. 中国兽医科学, 2010, 40(01): 45-50. [8] Qin YM. Preparation of Listeriolysin O monoclonal antibodies and preliminary deteetion of Listeria monocytogenes [D]. Harbin: Department of Preventive Veterinary Medicine, Northeast Agricultural University, 2007. (in Chinese) 秦玉敏. 李氏溶血素单克隆抗体制备及检测李氏溶血素的初步探索[D]. 哈尔滨: 东北农业大学预防兽医学系, 2007. [9] Yu KY, Noh Y, Chung M, et al. Use of monoclonal antibodies that recognize p60 for identification of Listeria monocytogenes [J]. Clin Diagn Lab Immunol, 2004, 11(3): 446-451.doi:10.1128/CDLI.11.3.446-451.2004
2015, Vol. 31 
