Richard Christelle, Drider Djamel, Fliss Ismael, Denery Sandra, Prevost Herve
Unité de Recherche sur la Qualité Microbiologique et Aromatique des Aliments (QM2A), ENITIAA, 44322 Nantes cedex 3, France.
Appl Environ Microbiol. 2004 Jan;70(1):248-54. doi: 10.1128/AEM.70.1.248-254.2004.
Polyclonal antibodies have been generated by immunization of rabbits with a chemically synthesized C-terminal part of divercin V41 (DvnCt) conjugated to the carrier protein keyhole limpet hemocyanin (KLH). The sensitivity and reactivity of the DvnCt-KLH-generated antibodies were evaluated by enzyme-linked immunosorbent assay (ELISA) using supernatant from cultures of 13 representative lactic acid bacterium strains, and specificity was confirmed by Western blot analysis. Anti-DvnCt-KLH antibodies were able to recognize not only divercin V41 but also enterocin P and piscicocin V1b, two other members of the class IIa bacteriocins. Production and activity of DvnV41 were evaluated by ELISA and activity tests during the growth of Carnobacterium divergens V41 in MRS medium containing or not containing Tween 80. Divercin V41, enterocin P, and piscicocin V1b were therefore purified by a single-step immunoaffinity chromatography method using a Sepharose matrix CNBr-activated directed binding of anti-DvnCt-KLH polyclonal antibodies.
通过用化学合成的、与载体蛋白钥孔血蓝蛋白（KLH）偶联的双歧菌素V41（DvnCt）C末端部分免疫兔子，制备了多克隆抗体。使用13种代表性乳酸菌菌株培养物的上清液，通过酶联免疫吸附测定（ELISA）评估了DvnCt-KLH产生的抗体的敏感性和反应性，并通过蛋白质印迹分析证实了其特异性。抗DvnCt-KLH抗体不仅能够识别双歧菌素V41，还能识别IIa类细菌素的另外两个成员肠球菌素P和鱼菌素V1b。在含有或不含吐温80的MRS培养基中培养分歧肉杆菌V41期间，通过ELISA和活性测试评估了双歧菌素V41的产生和活性。因此，使用琼脂糖基质CNBr活化的抗DvnCt-KLH多克隆抗体定向结合的单步免疫亲和色谱法，纯化了双歧菌素V41、肠球菌素P和鱼菌素V1b。
