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【目的】建立一种基于单克隆抗体的多元弧菌流式细胞仪检测技术。【方法】以副溶血弧菌表面蛋白r-OmpW的单克隆抗体(mAb)为基础,以细胞染色率为指标优化流式细胞仪检测副溶血弧菌时所需mAb的反应浓度和反应时间。通过菌落数对比评价在优化条件下流式细胞术方法的准确度、检出限和精密度。根据所建立的流式细胞术平台分析鉴定单抗对其它5种多元弧菌的特异性。【结果】流式细胞仪检测副溶血弧菌时所需r-OmpW单克隆抗体的优化反应浓度为20 mg/L,反应时间为60 min。当菌浓在104 107cells/mL范围时,检测值可信度较高,可特异性识别5种病原弧菌。对含不同菌体浓度的样品进行重复检测,变异系数均在7%以内。【结论】所建立的这种基于单克隆抗体的多元弧菌流式细胞仪检测技术可快速准确地检测多种病原弧菌。
【Objective】 To establish a multi-Vibrio flow cytometry assay based on monoclonal antibodies. 【Method】 Monoclonal antibody (mAb) of Vibrio parahaemolyticus surface protein r-OmpW was used as the basis to optimize the reaction concentration and reaction time of mAb when Vibrio parahaemolyticus was detected by flow cytometry. The accuracy, detection limit and precision of the flow cytometry method under optimal conditions were evaluated by comparison of the number of colonies. According to the establishment of flow cytometry platform to identify the identification of monoclonal antibodies against other five Vibrio species specificity. 【Result】 The optimal concentration of r-OmpW monoclonal antibody for Vibrio parahaemolyticus detected by flow cytometry was 20 mg / L and the reaction time was 60 min. When the bacteria concentration in 104 107cells / mL range, the detection of high reliability, can specifically identify five kinds of pathogens Vibrio. Samples containing different concentrations of bacteria were repeatedly tested, the coefficient of variation were within 7%. 【Conclusion】 The monoclonal antibody-based Vibrio versicolor flow cytometry was established to rapidly and accurately detect a variety of pathogenic Vibrio spp.