摘要
用E.coliO157∶H7菌体免疫BALB/c鼠,制备免疫脾细胞,与SP2/0骨髓瘤细胞融合,获得了16株分泌单克隆抗体（mAb）的杂交瘤细胞株。其中,1D3、2E7和2H7三株为O157∶H7特异单克隆抗体,Ig亚类分别为IgMκ、IgG2bκ和IgG2bκ,腹水抗体ELISA效价分别为103、105和106。用纯化的单克隆抗体2E7标记胶体金,制备金标抗体玻璃纤维素膜;将纯化的2H7和羊抗鼠IgG分别作为检测和对照捕捉抗体包被于硝酸纤维素膜（NC）上;组装成双抗夹心法O157∶H7胶体金免疫层析检测试纸条。用该试纸条可特异检测O157∶H7,敏感度为106CFU/mL,与相同单克隆抗体2H7和2E7建立的O157∶H7夹心ELISA检测方法的敏感度相当,试纸条简便快速,在1~5 min内可得出检测结果,便于O157∶H7的临床快速诊断与现场检测样品的快速筛查。
BALB/c mice were immunized with formaldehyde-killed E.coli O157∶H7 and their splenocytes were fused with myeloma cells SP2/0.As a result,16 hybridomas were generated and 3 of them（1D3,2E7 and 2H7） were identified to secret monoclonal antibodies（mAbs） to E.coli O157∶H7.The antibody isotypes were IgMκ（1D3）,IgG2bκ 92E7） and IgG2bκ（2H7）.The ELISA titers of ascitic fluids were 103,105 and 106,respectively.The gold particle in colloidal solution was coupled with purified monoclonal antibody 2E7,and then dispensed on the glass fibers.The purified monoclonal antibody 2H7 and goat anti-mouse IgG serving as capture antibody were coated separately on the nitrocellulose membrane as the test line（T line） and the control line（C line）.Then the sandwich-based gold immunochromatographic strip for the detection of E.coli O157∶H7 was assembled in regular sequence.The test results indicated that the strip was specific to O157∶H7 and the sensitivity was 106CFU/ml,which was equivalent to the sandwich ELISA established by using the same monoclonal antibodies 2E7 and 2H7.The strip is convenient and rapid for the clinical diagnosis and field investigation of E.coli O157∶H7 infection.
出处
《中国动物传染病学报》
CAS
2010年第4期47-53,共7页
Chinese Journal of Animal Infectious Diseases
基金
"十一五"国家科技支撑计划(2007BAD40B01)
作者简介
作者简介：夏兴霞，女，研究实习员，硕士研究生，主要从事病原微生物快速检测技术研究
