Padhye V V, Zhao T, Doyle M P
Department of Food Microbiology and Toxicology, University of Wisconsin-Madison 53706.
J Med Microbiol. 1989 Nov;30(3):219-26. doi: 10.1099/00222615-30-3-219.
Fourteen hybridoma cell lines were isolated that produced monoclonal antibodies (MAbs) to purified Verotoxins 1 and 2 (VT1, VT2) of Escherichia coli of serotype O157:H7. Of these MAbs, eight were obtained by immunisation of BALB/c mice with purified VT1, and six were obtained from BALB/c mice immunised with purified VT2. With the exception of MAb 1C5, with a heavy chain of IgG2b class, antibodies produced from mice immunised with heat-treated toxin were of IgM class. MAbs produced from mice immunised with heat-treated VT1 or VT2 reacted with both verotoxins in ELISA, and Western-blot analysis revealed that they reacted with subunit A and the A1 fragment of nicked subunit A of both toxins, but not with subunit B; furthermore, none of them neutralised Vero cytotoxicity or mouse lethality of either toxin. In contrast, MAbs produced from mice immunised with heat-treated and formalin-treated VT1 reacted in Western blots with subunits A and B of VT1 and subunit A, but not subunit B, of VT2, reacted in ELISA with VT1 only, and neutralised Vero cytotoxicity and mouse lethality of VT1 but not of VT2. Results indicate the existence of a common epitope on subunit A of VT1 and VT2 that is not responsible for the biological activity of these toxins, and that subunit B is essential for the biological activity of VT1. MAbs capable of reacting with both verotoxins from E. coli of serotype O157:H7 may be useful reagents for screening bacterial isolates capable of producing one or both of these toxins.
分离出了14种杂交瘤细胞系，它们产生针对血清型O157:H7大肠杆菌纯化的维罗毒素1和2（VT1、VT2）的单克隆抗体（MAb）。在这些单克隆抗体中，8种是通过用纯化的VT1免疫BALB/c小鼠获得的，6种是从用纯化的VT2免疫的BALB/c小鼠中获得的。除了重链为IgG2b类的单克隆抗体1C5外，用热处理毒素免疫小鼠产生的抗体均为IgM类。用热处理的VT1或VT2免疫小鼠产生的单克隆抗体在ELISA中与两种维罗毒素都发生反应，蛋白质印迹分析表明它们与两种毒素的亚基A和缺口亚基A的A1片段发生反应，但不与亚基B反应；此外，它们均未中和任何一种毒素的Vero细胞毒性或小鼠致死性。相比之下，用热处理和甲醛处理的VT1免疫小鼠产生的单克隆抗体在蛋白质印迹中与VT1的亚基A和B以及VT2的亚基A（但不与亚基B）发生反应，在ELISA中仅与VT1发生反应，并中和了VT1的Vero细胞毒性和小鼠致死性，但未中和VT2的。结果表明VT1和VT2的亚基A上存在一个共同表位，该表位与这些毒素的生物学活性无关，并且亚基B对VT1的生物学活性至关重要。能够与血清型O157:H7大肠杆菌的两种维罗毒素发生反应的单克隆抗体可能是筛选能够产生其中一种或两种毒素的细菌分离株的有用试剂。
