抗嗜肺军团菌血清8型单克隆抗体的制备及双抗体夹心酶联免疫吸附试验检测法的建立

Preparation of Monoclonal Antibodies Against Legionella pneumophila Serogroup 8 and Establishment of Antibody-Sandwich ELISA Method for Detecting this Pathogen

目的:制备针对嗜肺军团菌血清8型的单克隆抗体,并建立双抗体夹心酶联免疫吸附试验(ELISA)检测方法。方法:用甲醛灭活的嗜肺军团菌血清8型菌免疫BALB/c小鼠,采用杂交瘤技术制备抗嗜肺军团菌血清8型单克隆抗体,建立双抗夹心ELISA检测方法。结果:研制出8株能特异性分泌抗嗜肺军团菌血清8型单克隆抗体的杂交瘤细胞株,Ig类型分别为IgM(2株)、IgG3(1株)和IgG1(5株);利用IgG1型单抗6G10与6C7配对,建立了双抗夹心ELISA检测方法,该方法的最低检出浓度为2.6×105cfu/mL,除与金黄色葡萄球菌有微弱的交叉反应外,与14株其他血清型嗜肺军团菌、17株非嗜肺军团菌及11株非军团菌均无交叉反应,具有较高的特异性。结论:制备了具有高特异性和亲和力的抗嗜肺军团菌血清8型单克隆抗体,并建立了双抗夹心ELISA检测方法。

Objective： To preparate monoclonal antibodies（McAbs） anginst Legionella pneumophila serogroup 8 and establish an antibody-sandwich ELISA method for detection of the organism. Methods： BALB/c mice were immunized by formalin-killed L.pneumophila serogroup 8 and McAbs were prepared by using hybridoma technique. By using two McAbs, an antibody-sandwich ELISA was established. Results： Eight hybridomas producing antibodies against L.pneumophila serogroup 8 were obtained. Ig isotypes of the McAbs were IgM, IgG3 and IgG1. An antibody-sandwich ELISA was established and the sensitivity for sonicated cells was about 2.6x105 cfu/mL. No cross-reactivity was detectable in 14 other serogroup L.pneumophila, 17 non-pneumophila legionellae and 11 other bacteria, except Staphylococcus aureus which has a weak cross reaction. Conclusion： Two hybridomas producing high specificity and affinity monoclonal antibodies against L. pneumophila serogroup 8 can provide for antibody-sandwich ELISA.