6种沙门菌单克隆抗体的制备和效能评价

Preparation and Evaluation of Monoclonal Antibodies Against Six Salmonella Pathogens

目的:制备稳定、特异、高亲和性的分别针对甲型副伤寒沙门菌、乙型副伤寒沙门菌、丙型副伤寒沙门菌、肠炎沙门菌、伤寒沙门菌和猪霍乱沙门菌的单克隆抗体。方法:用甲醛灭活的菌液抗原免疫BALB/c小鼠,取脾细胞与SP2/0骨髓瘤细胞融合;用灭活的菌液包被酶标板,ELISA筛选阳性克隆株,建立细胞系;选取高效分泌杂交瘤细胞,常规制备腹水并纯化,进行单抗特异性与亲和性评价。结果:筛选得到分泌6种沙门菌相应单克隆抗体的杂交瘤细胞株,获得高亲和性单抗;所有单抗与大部分病原菌(包括7种沙门菌、3株志贺菌、2株李斯特菌、4株致病性大肠杆菌、2株霍乱弧菌)无交叉反应,但由于同类型O抗原的广泛分布,抗乙型副伤寒沙门菌单抗与鼠伤寒沙门菌、抗伤寒沙门菌单抗与肠炎沙门菌有明显的交叉反应。结论:沙门菌单抗的制备,为感染性腹泻的监测、诊断奠定了基础。

Objective： To prepare six stable, specific monoclonal antibodies（mAbs） with high affinity against Sal- monella paratyphi A, S.paratyphi B, S.paratyphi C, S.enteritidis, S.typhi and S.choleraesuis respectively. Methods： BALB/c mice were immunized with bacteria inactivated by formaldehyde, and then myeloma cell SP2/0 were fused with splenocyte which taken from immunized BALB/c mice. The positive hybridomas were screened with inactive pathogens, and then cell lines were established to produce mAbs. Purified mAbs were evaluated by the method of indirect ELISA. Results： Hybridomas cell lines related to six Salmonella pathogens respectively were established and mAbs with high affinity were obtained. All of mAbs did not cross-react with a wide range of food-borne pathogens, including Salmonella spp., Shigella spp., Listeria spp., Escherichia coli spp., and Vibrio cholera, but mAbs against S.paratyphi B reacted with S.typhimurium and mAbs against S.typhi reacted with S.enteritidis for their identical O-antigens. Conclusion： Six mAbs against Salmonella pathogens will contribute to diagnosis and sci- entific research of diarrhea.