Genetic Engineering Interdisciplinary Program, Graduate School, Kasetsart University, 50 Ngam Wong Wan Road, Chatuchak, Bangkok, 10900, Thailand.
Department of Research Affairs, Faculty of Dentistry, Chulalongkorn University, Bangkok, 10330, Thailand.
Anal Bioanal Chem. 2024 Dec;416(30):7357-7368. doi: 10.1007/s00216-024-05576-3. Epub 2024 Oct 11.
PCV2 is a significant epidemic agricultural pathogen that causes a variety of swine diseases. PCV2 infections have significant economic impact on the swine industry, making effective strategies for rapid detection of PCV2 in pigs essential. Herein, we report on the synthesis of the so-called nano-MIPs which can be utilized for molecular recognition of PCV2. The morphology and structure of nano-MIPs were characterized using scanning electron microscopy (SEM). Nano-MIPs are spherical with sizes around 120-150 nm. Binding experiments demonstrate that the fluorescence intensity of PCV2 samples decreases proportionally to increasing the concentration of nano-MIPs due to quenching, while non-imprinted polymer nanoparticles (nano-NIPs) do not affect the signal. The Stern-Volmer constant of nano-MIPs binding to PCV2 was 1.3 × 10 mL/µg, whereas nano-NIPs led to 7 × 10 mL/µg, i.e., 1.8 orders of magnitude lower. The detection limit for binding MIP particles to PCV2 by fluorescence measurements is 47 µg/mL. This affinity test allows for designing both direct and competitive quartz crystal microbalance (QCM) assays for PCV2 leading to QCM measurements. The QCM results show nano-MIPs binding to PCV2 immobilized on the sensor surface with appreciable reproducibility. QCM sensor characteristics reveal signal saturation above around 200 µg/mL at a response of - 354 Hz and an LOD of approximately 35 µg/mL. Nano-MIPs also show selectivity factors of 2-5 for CSFV and PRRSV probably because the three viruses have similar diameters around 50 nm.
PCV2 是一种重要的流行农业病原体，可引起多种猪病。PCV2 感染对养猪业有重大的经济影响，因此，快速检测猪 PCV2 的有效策略至关重要。在此，我们报告了所谓的纳米 MIPs 的合成，该 MIPs 可用于 PCV2 的分子识别。使用扫描电子显微镜（SEM）对纳米 MIPs 的形态和结构进行了表征。纳米 MIPs 呈球形，尺寸约为 120-150nm。结合实验表明，由于猝灭，PCV2 样品的荧光强度与纳米 MIPs 浓度的增加成正比降低，而非印迹聚合物纳米颗粒（nano-NIPs）则不会影响信号。纳米 MIPs 与 PCV2 结合的 Stern-Volmer 常数为 1.3×10mL/µg，而 nano-NIPs 则导致 7×10mL/µg，即低 1.8 个数量级。通过荧光测量检测到纳米 MIP 颗粒与 PCV2 结合的检测限为 47µg/mL。该亲和力测试允许设计直接和竞争石英晶体微天平（QCM）分析用于 PCV2 的 QCM 测量。QCM 结果表明，纳米 MIPs 与固定在传感器表面的 PCV2 结合具有相当的重现性。QCM 传感器特性显示，在响应值为-354Hz 时，信号饱和约为 200µg/mL，LOD 约为 35µg/mL。纳米 MIPs 对 CSFV 和 PRRSV 的选择性因子为 2-5，可能是因为这三种病毒的直径约为 50nm，相似。
