Qian Hongliang, Pang Ervinna, Du Qingyun, Chang Jason, Dong Jin, Toh Say Ling, Ng Fook Kheong, Tan Ai Ling, Kwang Jimmy
Animal Health Biotechnology Group, Temasek Life Sciences Laboratory, 1 Research Link, National University of Singapore, Singapore 117604, Republic of Singapore.
Appl Environ Microbiol. 2008 Feb;74(3):833-9. doi: 10.1128/AEM.01559-07. Epub 2007 Dec 7.
Campylobacter species are important enteric pathogens causing disease in humans and animals. There is a lack of a good immunological test that can be used routinely to separate Campylobacter jejuni from other Campylobacter species. We produced monoclonal antibodies (MAbs) directed against the major outer membrane protein (MOMP) of C. jejuni using recombinant MOMP as the antigen. One MAb, designated MAb5C4 and of the immunoglobulin G1 isotype, was found to be potentially specific for C. jejuni. Dot blots demonstrated that MAb5C4 reacted with all 29 isolates of C. jejuni tested but did not react with 2 C. jejuni isolates, 26 other Campylobacter spp. isolates, and 19 non-Campylobacter isolates. Western blotting showed that MAb5C4 bound to a single protein band approximately 43 kDa in size, corresponding to the expected size of C. jejuni MOMP. The detection limit of MAb5C4 in a dot blot assay was determined to be about 5 x 10(3) bacteria. The epitope on the MOMP was mapped to a region six amino acids in length with the sequence 216GGQFNP221, which is 97% conserved among C. jejuni strains but divergent in other Campylobacter spp.; a GenBank search indicated that 95% of C. jejuni isolates will be able to be detected from non-Campylobacter spp. based on the highly specific and conserved region of the GGQFNP polypeptide. The epitope is predicted to be located in a region that is exposed to the periplasm. MAb5C4 is a potentially specific and sensitive MAb that can be used for the specific detection and identification of C. jejuni.
弯曲杆菌属是引起人和动物疾病的重要肠道病原体。目前缺乏一种可常规用于将空肠弯曲菌与其他弯曲杆菌属物种区分开来的良好免疫学检测方法。我们使用重组主要外膜蛋白（MOMP）作为抗原，制备了针对空肠弯曲菌主要外膜蛋白的单克隆抗体（MAb）。发现一种单克隆抗体，命名为MAb5C4，属于免疫球蛋白G1同种型，可能对空肠弯曲菌具有特异性。斑点印迹表明，MAb5C4与所有测试的29株空肠弯曲菌分离株发生反应，但不与2株空肠弯曲菌分离株、26株其他弯曲杆菌属物种分离株以及19株非弯曲杆菌分离株发生反应。蛋白质印迹显示，MAb5C4与一条大小约为43 kDa的单一蛋白带结合，这与空肠弯曲菌MOMP的预期大小相对应。在斑点印迹分析中，MAb5C4的检测限确定为约5×10³个细菌。MOMP上的表位被定位到一个长度为6个氨基酸的区域，序列为216GGQFNP221，该序列在空肠弯曲菌菌株中97%保守，但在其他弯曲杆菌属物种中存在差异；GenBank搜索表明，基于GGQFNP多肽的高度特异性和保守区域，95%的空肠弯曲菌分离株能够从非弯曲杆菌属物种中检测出来。预测该表位位于暴露于周质的区域。MAb5C4是一种潜在的特异性和敏感性单克隆抗体，可用于空肠弯曲菌的特异性检测和鉴定。
