基于近红外免疫层析技术的食源性副溶血弧菌快速检测方法研究

Rapid Detection of Foodborne Vibrio parahaemolyticus, Based on a Near-infrared Immunochromatographic Technique

建立一种应用近红外荧光免疫层析技术快速检测副溶血弧菌的方法。采用近红外荧光标记副溶血弧菌单克隆抗体,将副溶血弧菌多克隆抗体和羊抗鼠IgG多克隆抗体喷涂于硝酸纤维膜上分别作为检测线和质控线,研制检测副溶血弧菌的近红外免疫层析试纸条及配套标准物质。研究表明,所建立的近红外免疫法对检测副溶血弧菌具有良好的特异性和较高的灵敏度,最低检测限为1.2×10~2 CFU/mL,与沙门氏菌、金黄色葡萄球菌、大肠埃希氏菌、单增李斯特菌无交叉反应。通过效果评价试验发现,与传统检测方法相比较,所建立的近红外荧光法检测时间时间最短,检测限与RT-PCR法接近。近红外荧光法在45 min内即可完成检测,可用于食品中副溶血弧菌的高效检测,为食品安全监管提供可靠的技术支持。

This study was designed to establish a method for the rapid detection of Fibrio parahaemolyticus, using a near-infrared fluorescence immunochromatographic technique. Near-infrared fluorescence was used to label anti-Vibrio parahaemolyticus monoclonal and polyclonal antibodies, and a goat anti-mouse IgG polyclonal antibody. These were then coated onto a nitrocellulose membrane as the detection line and control line, respectively, to develop a near-infrared immunochromatographic test dipstick for the detection of Vibrio parahaemolyticus and a standard substance. The results showed that the established near-infrared immunochromatographic method had good specificity and high sensitivity for the detection of Vibrio parahaemolyticus, the limit of detection was 1.2×10^2 CFU/mL, and there were no cross-reactions with Salmonella, Staphylococcus aureus, Escherichia coli, or Listeria monocytogenes. The performance evaluation test found that, compared with the traditional detection method, the established near-infrared fluorescence method had a shorter detection time, and a detection limit close to that of the real-time polymerase chain reaction （RT-PCR） method. A measurement using this near-infrared immunochromatographic method could be completed in 45 minutes. This method can be used for the efficient detection of Vibrio parahaemolyticus in food and provide reliable technical support for food safety supervision.