摘要
从已建立的抗沙门氏菌单抗中筛选出适用于肠炎沙门氏菌ELISA检验的3-47-26单抗,采用筛选强阳性杂交瘤细胞株制备了高效价的3-47-26单抗。改进用辣根过氧化物酶标记高效价单抗的方法,进行了HRP标记单抗的免疫生物学特性的鉴定。确定了包被浓度和酶标抗体的工作浓度:HRP-3-47-26为1∶100;LPS的包被浓度为400ng/mL。试验中采用LPS与多聚赖氨酸的结合,解决了包被过程中的解吸附作用,增强了包被的稳定性,同时也减少了假阳性反应,优化了包被过程。建立了检测肠炎沙门氏菌的抗原竞争ELISA方法,利用样品中的LPS抑制酶标抗体与包被的LPS结合,以降低底物反应的颜色从而达到检测的目的。通过对210份肠炎沙门氏菌感染鸡泄殖腔棉拭子、羽毛和组织样品先筛选后鉴定的方法进行检测具有明显的抑制作用,通过与国标法对大量的样品检测结果比较表明,竞争ELISA方法的检出率为18.09%;检出阳性样品36份,国标法检出率为17.14%;两者符合率为97.14%。竞争ELISA敏感性和特异性分别为94.4%和97.7%。从而为肠炎沙门氏菌的检测提供了一种敏感、特异的检测方法。
A monoclonal antibody （McAb） 3-47-26 specific to O9,was used to establish competitive Enzyme -finked irnmunosorbed assays（C-ELISA） of-detection for Salmonella enteritidis.A hybridomas cell strain 3 47 26 was screened and prepared the high valence monoclonal antibody. The linking method of McAb with HRP was improved and the specificity of HRP-3-47-26 was accredited. The concentration of coating of antigen LPS and HRP-3-47-26 was determined ： HRP-3-47-26, 1 ： 100; LPS,400 ng/mL. Stable coating and minimal false positives were achieved by conjugating LPS to poly-L-lysine, which solved the desorption problem. S.enteritidis LPS in samples competed with S.enteritidis LPS coated on microtitre plate, and competition first reduced binding of McAb to the S.enteritidis LPS on the plate hence to reduce the chromogenic signal, which the aim of detection for sample. The C-ELISA was very specific and no cross reaction, and it was observed in detection sera induced by related Sanunonella A F, compared with routine detective method for the detection of S.enteritidis, and the sensitivity and specificity of the C-ELISA were 94.4 % and 97.7 % respectively.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2006年第2期196-200,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
山西省科技厅项目(项目编号:022044)
关键词
肠炎沙门氏菌
单抗
检测
Samonella enteritidis
monoclonal antibody
detection
作者简介
黄素珍（1955-）女，教授，从事预防兽医学研究．
