基于磁分离-上转换荧光标记的副溶血性弧菌免疫检测新方法研究

Novel Immunodetection Method for Vibrio parahaemolyticus Based on Magnetic Separation and Upconversion Nanoparticles Labeling

DOI：10.3969/j.issn.1673-1689.2014.07.002

中文关键词: 副溶血性弧菌 上转换荧光 磁分离 FlaA抗体

英文关键词: Vibrio parahaemolyticus, upconversion fluorescence, magnetic separation, antibodies of flagellin

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中文摘要:

将上转换荧光标记与磁分离富集技术相结合，通过免疫识别成功构建了一种检测副溶血性弧菌（Vibrio parahaemolyticus）的新方法。首先通过水/溶剂热法合成NaY0.78F4：Yb0.20，Er0.02上转换荧光纳米颗粒，并对其表面进行氨基功能化修饰，同时一步法合成了氨基化Fe3O4磁性纳米颗粒。分别将副溶血性弧菌全菌抗体与磁性纳米颗粒连接作为捕获探针，将副溶血性弧菌的鞭毛蛋白A抗体与上转换荧光纳米颗粒连接作为信号探针。通过免疫识别形成捕获探针-目标菌-显示探针的"三明治"结构复合物，利用980 nm激光诱导荧光进行检测。在实验优化条件下，上转换荧光强度与副溶血性弧菌浓度在5×103~5×105 cfu/mL范围内呈良好线性关系，检测限为1×103 cfu/mL。用鲫鱼进行加标回收实验，结果表明，本方法准确性良好。

英文摘要:

Based on upconversion fluorescence labeling, magnetic separation and immunological recognition, a novel detection method of Vibrio parahaemolyticus was constructed. Primarily, the NaY0.78F4: Yb0.20, Er0.02 upconversion nanoparticles(UCNPs) were synthesized through hydrothermal method, and then amino group was modified on the surface of UCNPs. Meanwhile, amino-modified MNPs were synthesized by one-step process. Secondly, the antibodies of Vibrio parahaemolyticus binding with MNPs and antibodies of flagellin binding with UCNPs were used as capture probes and signal probes, respectively. The composite of MNPs-target bacteria-UCNPs was obtain, and detected by 980 nm laser. Under the optimum conditions, the upconversion fluorescence was a good linear in the range of 5×103~5×105 cfu/mL of Vibrio parahaemolyticus, and the detection of limit was 1×103 cfu/mL. This method was accurate in the recovery text in the crucian.

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