Rapid direct determination using combined separation by prepared immunomagnetic and flow cytometry of Flavobacterium psychrophilum
- 作者：Kyoko Hibi ; Kohji Mitsubayashi ; Hideo Fukuda ; Hideki Ushio ; Tetsuhito Hayashi ; Huifeng Ren ; Hideaki Endo
- 关键词：Immunomagnetic separation ; Flow cytometry ; Iron particles ; Flavobacterium psychrophilum
- 刊名：Biosensors and Bioelectronics
- 出版年：2007
- 期刊代码：12_09565663
- 类别：ch
- 出版时间：15 April 2007
- 卷：22
- 期：9-10
- 页码：1916-1919
- 文件大小：202 K
摘要
Flavobacterium psychrophilum, the causative agent of bacterial cold-water disease (BCWD), was originally isolated from coho salmon Oncorhychus kisutch in the USA. Bacterial cold-water disease has since been spreading throughout Japan and has caused serious damage to populations of ayu Plecoglossus altivel in many farms and rivers. The rapid method of detecting for F. psuchrophilum is requested, however, traditional methods are laborious because of complicated assay procedures. In this study, a rapid method of detecting F. psychrophilum was developed using a modified method of flow cytometry (FCM) analysis and immunomagnetic separation (IMS). Magnetic iron, in small particles, was prepared by the reaction of a mixture of ferric and ferrous ions under alkaline conditions. The particles were coated with antiserum against F. psychrophilum by dextran. Polyclonal antibodies (anti-F. psychrophilum) conjugated with fluorescein isothiocyanate (FITC) were reacted with F. psychrophilum, and then prepared immunomagnetic were applied using IMS, followed by FCM determination. A good correlation was observed between the cell numbers determined by the FCM method and the traditional method in the range of 102–108 cells ml−1. The FCM analysis could count cells within 1 min, and the total analysis time, including sample preparation, was less than 2 h.