摘要
为制备抗布鲁氏菌单克隆抗体(MAb),本研究用灭活的布鲁氏菌疫苗M5-90株免疫BALB/c小鼠,利用细胞融合技术获得杂交瘤细胞。以布氏杆菌疫苗株M5-90、S-19及小肠结肠炎耶尔森菌O∶9的脂多糖(LPS)分别作为抗原包被酶标板,对杂交瘤细胞进行筛选,获得两株稳定分泌抗光滑型布氏杆菌LPS(S-LPS)MAb的细胞4G6和16C5。特异性分析表明,MAb 4G6除与耶尔森O∶9全菌体有轻微的交叉反应外与其它革兰氏阴性菌无交叉,而16C5完全排除了与其他各菌的交叉反应。Ig亚型分析表明,所制备MAb的轻链亚类均为κ,4G6重链为IgM,16C5重链为IgG3。用Protein G亲和层析纯化MAb 16C5腹水并初步用于竞争ELISA方法,检测布氏杆菌抗体水平,结果表明了该实验方法的有效性。本研究制备的MAb 16C5具有高度的特异性和敏感性,排除了与其他革兰氏阴性菌的交叉反应,为建立一种诊断布鲁氏病的快速、有效、敏感的方法奠定了有力的基础。
To preparation monoclonal antibody （MAb） against smooth Brucella LPS, the B.mefitensis M5-90 were inactivated and used to immunize BALB/c mice, Hybridomas were obtained by standard cell fusion techniques. Two cell strains secreting MAb against S-LPS, named 4G6 and 16C5, were obtained by Ⅰ-ELISA screening with Brucella vaccine strains M5-90, S-19 and Y. enterocolitica 0：9 as antigens. MAb 16C5 was highly specific and had no cross reaction with all Gram-negative bacteria tested, while MAb 4G6 had no cross reaction with all but Y.enterocolitica 0：9. MAb 4G6 belonged to IgM while MAb 16C5 belonged to IgG3, and both light chains belonged to rc. Ascites of MAb 16C5 was purified by protein G and successfully used in competitive ELISA to detect antibodies against Brucella in cattle serum. The highly sensitivee and specific MAb 16C5 would provide a potential agent for diagnosis of brucellosis.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2009年第8期642-645,649,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
国家863攻关(20060102Z449)
