A. N. Bach Institute of Biochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Leninsky Prospect 33, 119071 Moscow, Russia.
Department of Chemical Enzymology, Faculty of Chemistry, M. V. Lomonosov Moscow State University, Leninskie Gory 1, 119991 Moscow, Russia.
Biosensors (Basel). 2023 Apr 16;13(4):477. doi: 10.3390/bios13040477.
In this study, a homogeneous fluorescence polarization immunoassay (FPIA) for the detection of hazardous aquatic toxin okadaic acid (OA) contaminating environmental waters was for the first time developed. A conjugate of the analyte with a fluorophore based on a fluorescein derivative (tracer) was synthesized, and its interaction with specific anti-OA monoclonal antibodies (MAbs) was tested. A MAbs-tracer pair demonstrated highly affine immune binding ( = 0.8 nM). Under optimal conditions, the limit of OA detection in the FPIA was 0.08 ng/mL (0.1 nM), and the working range of detectable concentrations was 0.4-72.5 ng/mL (0.5-90 nM). The developed FPIA was approbated for the determination of OA in real matrices: river water and seawater samples. No matrix effect of water was observed; therefore, no sample preparation was required before analysis. Due to this factor, the entire analytical procedure took less than 10 min. Using a compact portable fluorescence polarization analyzer enables the on-site testing of water samples. The developed analysis is very fast, easy to operate, and sensitive and can be extended to the determination of other aquatic toxins or low-molecular-weight water or food contaminants.
本研究首次开发了一种用于检测环境水中有害水生毒素 okadaic 酸（OA）的均相荧光偏振免疫分析（FPIA）。合成了一种基于荧光素衍生物（示踪剂）的分析物与荧光团的缀合物，并测试了其与特异性抗 OA 单克隆抗体（MAb）的相互作用。MAb-示踪剂对表现出高度亲和免疫结合（=0.8 nM）。在最佳条件下，FPIA 中 OA 的检测限为 0.08 ng/mL（0.1 nM），可检测浓度的工作范围为 0.4-72.5 ng/mL（0.5-90 nM）。开发的 FPIA 已批准用于测定实际基质中的 OA：河水和海水样本。未观察到水的基质效应；因此，在分析之前不需要进行样品制备。由于这个因素，整个分析过程耗时不到 10 分钟。使用紧凑型便携式荧光偏振分析仪可实现现场测试水样。所开发的分析方法快速、易于操作且灵敏，可扩展用于测定其他水生毒素或低分子量水或食品污染物。
