应用Dot-ELISA快速检测牙鲆腹水病病原的研究

Rapid Detection of Pathogenic Bacteria of Ascites in Japanese Flounder(Paralichthys olivaceus)by Dot-ELISA

选用硝酸纤维素膜作固相载体,建立检测迟钝爱德华氏菌的斑点酶联免疫吸附试验诊断方法。根据棋盘试验,确定迟钝爱德华氏菌免疫血清最佳工作浓度为1∶800,酶标抗体最佳工作浓度为1∶200,以出现明显清晰斑点者判定为阳性。用该方法检测时,迟钝爱德华氏菌呈阳性,温和气单胞菌、嗜水气单胞菌、河流弧菌、溶藻弧菌、鳗弧菌、腐败希瓦氏菌、产碱普罗威斯登菌、阪崎肠杆菌和大肠杆菌均呈阴性。试验结果表明,Dot-ELISA方法简便、特异、快速、结果直观,便于在基层推广使用。

A Dot-Enzyme Linked Immunosorbent Assay（Dot-ELISA）was successfully established for detection of Edwardsiella tarda found in Japanese flounder（Paralichthys olivaceus）by using nitrocellulose membrane as solid carrier.The working concentration of serum samples was 1∶800 dilution and that of the enzyme-labeled goat anti-rabbit IgG was 1∶200 dilution by chessboard assay.The samples with clear and obvious dots were judged as positive reaction.Edwardsiella tarda was positive in the Dot-ELISA,while Aeromonas sobria,A.hydrophila,Vibrio fluvialis,V.alginolyticus,V.anguillarum,Shewanella putrefaciens,Providencia alcalifaciens,Enterobacter sakazakii and Escherichia coli were negative in the assay.The trial indicated that this method was simple,specific,quick,direct,easy extention and application at basic units.