Bentley A T, Klebba P E
Department of Biological Sciences, University of Notre Dame, Indiana 46556.
J Bacteriol. 1988 Mar;170(3):1063-8. doi: 10.1128/jb.170.3.1063-1068.1988.
We studied the reactivity of 66 anti-Escherichia coli B/r porin monoclonal antibodies (MAbs) with several E. coli and Salmonella typhimurium strains. Western immunoblots showed complete immunological cross-reactivity between E. coli B/r and K-12; among 34 MAbs which recognized porin in immunoblots of denatured outer membranes of E. coli B/r, all reacted with OmpF in denatured outer membranes of E. coli K-12. Extensive reactivity, although less than that for strain B/r (31 of 34 MAbs), occurred for porin from a wild-type isolate, E. coli O8:K27. Only one of the MAbs reacted with porin in denatured outer membranes of S. typhimurium. Even with immunochemical amplification of the Western immunoblot technique, only six MAbs recognized S. typhimurium porin (OmpD), demonstrating that there is significant immunological divergence between the porins of these species. Antibody binding to the bacterial surface, which was analyzed by cytofluorimetry, was strongly influenced by lipopolysaccharide (LPS) structure. An intact O antigen, as in E. coli O8:K27, blocked adsorption of all 20 MAbs in the test panel. rfa+ E. coli K-12, without an O antigen but with an intact LPS core, bound seven MAbs. When assayed against a series of rfa E. coli K-12 mutants, the number of MAbs that recognized porin surface epitopes increased sequentially as the LPS core became shorter. A total of 17 MAbs bound porin in a deep rough rfaD strain. Similar results were obtained with S. typhimurium. None of the anti-E. coli B/r porin MAbs adsorbed to a smooth strain, but three antibodies recognized porin on deep rough (rfaF, rfaE) mutants. These data define six distinct porin surface epitopes that are shielded by LPS from reaction with antibodies.
我们研究了66种抗大肠杆菌B/r孔蛋白单克隆抗体（MAb）与几种大肠杆菌和鼠伤寒沙门氏菌菌株的反应性。蛋白质免疫印迹显示大肠杆菌B/r和K-12之间存在完全的免疫交叉反应性；在34种能在大肠杆菌B/r变性外膜免疫印迹中识别孔蛋白的MAb中，所有抗体都能与大肠杆菌K-12变性外膜中的OmpF发生反应。来自野生型分离株大肠杆菌O8:K27的孔蛋白也出现了广泛的反应性，尽管不如B/r菌株（34种MAb中的31种）。只有一种MAb能与鼠伤寒沙门氏菌变性外膜中的孔蛋白发生反应。即使采用蛋白质免疫印迹技术的免疫化学放大方法，也只有六种MAb能识别鼠伤寒沙门氏菌孔蛋白（OmpD），这表明这些物种的孔蛋白之间存在显著的免疫差异。通过细胞荧光测定法分析的抗体与细菌表面的结合受到脂多糖（LPS）结构的强烈影响。完整的O抗原，如在大肠杆菌O8:K27中，会阻断测试组中所有20种MAb的吸附。rfa+大肠杆菌K-12没有O抗原，但有完整的LPS核心，能结合7种MAb。当针对一系列rfa大肠杆菌K-12突变体进行检测时，随着LPS核心变短，识别孔蛋白表面表位的MAb数量依次增加。在深度粗糙的rfaD菌株中，共有17种MAb能结合孔蛋白。鼠伤寒沙门氏菌也得到了类似结果。没有一种抗大肠杆菌B/r孔蛋白的MAb能吸附到光滑菌株上，但有三种抗体能识别深度粗糙（rfaF、rfaE）突变体上的孔蛋白。这些数据确定了六种不同的孔蛋白表面表位，它们被LPS屏蔽而不与抗体发生反应。
