Chibani Azaïez S R, Fliss I, Simard R E, Moineau S
Department of Biochemistry and Groupe de Recherche en Ecologie Buccale (GREB), Faculté de Médecine Dentaire, Université Laval, Québec, Canada G1K 7P4.
Appl Environ Microbiol. 1998 Nov;64(11):4255-9. doi: 10.1128/AEM.64.11.4255-4259.1998.
Phage Q38, a representative member of the c2 species, was purified by CsCl gradient and used to immunize BALB/c mice. Monoclonal antibodies (MAbs) were raised and then characterized by enzyme-linked immunosorbent assay. Two MAbs of isotype immunoglobulin G2a, designated 2A5 and 6G7, reacted only with phages belonging to the c2 species and not with phages of the 936 and P335 species, with a Lactococcus lactis cell extract, or with phage DNA. Immunoelectron microscopy showed that both MAbs recognized only phage head proteins. They did not react with any denatured phage proteins in Western blot assays. However, when the nitrocellulose membranes were treated with a Triton-based buffer to assist in protein renaturation, MAbs 2A5 and 6G7 recognized the two major capsid proteins with molecular masses of 80 and 170 kDa. Competitive inhibition tests showed that the two MAbs bind to overlapping epitopes. These MAbs may be a useful tool for monitoring c2 bacteriophages during dairy fermentation and in genetic studies.
噬菌体Q38是c2种的代表性成员，通过氯化铯梯度离心进行纯化，并用于免疫BALB/c小鼠。制备了单克隆抗体（MAb），然后通过酶联免疫吸附测定进行表征。两种免疫球蛋白G2a同型的单克隆抗体，命名为2A5和6G7，仅与属于c2种的噬菌体发生反应，而不与936和P335种的噬菌体、乳酸乳球菌细胞提取物或噬菌体DNA发生反应。免疫电子显微镜显示，两种单克隆抗体都只识别噬菌体头部蛋白。在蛋白质印迹分析中，它们不与任何变性的噬菌体蛋白发生反应。然而，当用基于 Triton 的缓冲液处理硝酸纤维素膜以辅助蛋白质复性时，单克隆抗体 2A5 和 6G7 识别出分子量分别为 80 kDa 和 170 kDa 的两种主要衣壳蛋白。竞争性抑制试验表明，这两种单克隆抗体结合重叠表位。这些单克隆抗体可能是在乳制品发酵过程中和基因研究中监测c2噬菌体的有用工具。
