牙鲆腹水病病原免疫血清的制备和纯化试验研究

Preparation and Purification of Pathogenic Bacterial Immune Serum from Japanese Flounder(Paralichthys olivaceus) with Ascites

用迟钝爱德华氏菌免疫家兔,制备出高效价迟钝爱德华氏菌免疫血清。先制备迟钝爱德华氏菌抗原,耳缘静脉注射免疫家兔。经微量反应板法检测血清效价,效价达到1∶2560;用Millipore Montage抗体纯化试剂纯化抗体,纯化的抗体经SDS-PAGE电泳,杂带较少,条带主要集中于50 kD处;用斑点酶联免疫吸附试验检测时,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,特异、高效价的迟钝爱德华氏菌免疫血清,为快速检测牙鲆腹水病病原奠定了基础。

The methods used in preparation of high-titer anti-Edwardsieua tarda（Et） serum by immunized rabbits with E.trada were described as the following：Firstly,the antigen of Et was prepared by the rabbits vaccinated by an ear vein.The agglutination efficiency of the immune serum was found to be 1︰2560 with micro-reaction plate.Then,the antibody was purified by Millipore Montage antibody purified kit,and the purified antibodies by SDS-PAGE electrophoresis with less miscellaneous focus had the main strip of about 50 kD.E.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 specificity and high titer of immune serum were produced in this study,leading to a strategy to detect Bifidobacteriumin dairy products serologically.The specificity and high titer E.tarda immune serum provide the foundation for the rapid detection of pathogenic bacteria in Japanese flounder（Paralichthys olivaceus）with ascites.