光热效应驱动的纸芯片可视化检测前列腺特异性抗原的研究

Paper-based analytical device (PAD) has been the subject of great interest for Point-Of-Care (POC) detection of tumor biomarkers, such as Prostate specific antigen (PSA). However, a number of current PADs still have to rely on complex detection equipment or image-analytical processes for quantitative readout, which limits their practical operability. Photothermal effect has been widely applied in photothermal therapy, but rarely investigated in biomedical sensing, especially in PAD. In our previous study, the photothermal effect of Prussian blue nanoparticles (PB NPs) had been applied in conventional immunoassay of PSA. Herein, by fabricating a PSA immuno-detection system on PAD, we plan to develop a photothermal effect-driven PAD for simple, rapid and visual quantitative detection of PSA. PB NPs generated from the immuno-detection system, poly (N-isopropylacrylamide) hydrogel and the on-chip moving distance of the hydrogel-released dye are proposed to act as the photothermal agent, thermal-responsive sensing material and naked-eye visual quantification signal, respectively. The driving mechanism of dye-release from the hydrogel by the PB NPs-based photothermal effect would be revealed. We also plan to evaluate the PSA detection performance of the PAD. In addition, the accuracy of the PAD for PSA determination in practical serum samples would be validated. Therefore, this project is expected to provide both significant theoretical research foundation and experimental basis for application of photothermal effect in PADs for tumor biomarker detection.

纸芯片在前列腺特异性抗原（PSA）等肿瘤标志物的现场即时（POC）检测中备受关注，但目前常见纸芯片仍然借助复杂检测仪器或图片分析程序进行定量读数，实际操作不便捷。光热效应被广泛用于光热治疗，但在生物医学传感尤其在纸芯片中研究尚少。我们在前期研究中已将普鲁士蓝纳米粒子（PB NPs）的光热效应应用到PSA的常规免疫检测中。本项目通过在纸芯片上构建PSA免疫检测体系，采用检测体系中生成的PB NPs为光热剂，以聚―(N-异丙基丙烯酰胺)水凝胶为热敏传感材料，以光热效应驱动水凝胶所释放染料的移动距离为肉眼可视定量检测指标，从而研究一种基于光热效应驱动的简单、快速、肉眼可视化PSA定量检测纸芯片。本项目将揭示纸芯片上光热效应对水凝胶定量释放染料的驱动机理，阐明其检测性能，证实其用于实际血清样本检测的准确度，为光热效应在肿瘤标志物检测纸芯片中的应用提供理论基础和实验依据。

纸芯片被广泛应用于前列腺特异性抗原（PSA）等肿瘤标志物的免疫检测，但目前大量纸芯片仍然依赖复杂分析仪器输出定量检测信号。近年来，光热效应在生物传感尤其现场即时检测中显现出较大的应用潜力。本项目将光热效应引入到免疫检测纸芯片中，借助普鲁士蓝纳米（PB NPs）光热剂和聚（N–异丙基丙烯酰胺）—丙烯酰胺（NIPAM–AcAm）热敏水凝胶，研究了一种基于光热效应驱动的定量信号肉眼可视化PSA免疫检测芯片。主要研究包括：（1） 采用掺杂氧化石墨烯（GO）的NIPAM–AcAm水凝胶为光热响应材料，利用热成像跟踪芯片界面上的热信号，探讨了芯片界面上光热效应驱动水凝胶的相转变行为及其定量泵送染料的工作机制；（2）采用合成PB的NIPAM–AcAm水凝胶为光热传感材料，构建了一种纸芯片光热传感平台，验证了芯片界面上光热效应驱动水凝胶所释放染料的移动距离以及热成像作为定量检测信号的可行性；（3）在上述研究基础上，设计了具有夹—仓嵌入式组装结构的纸基—聚甲基丙烯酸甲酯（PMMA）光热免疫检测芯片。将芯片（纸基底）免疫反应体系中捕获的Fe3O4 NPs转换为比色/光热双探针—PB NPs，通过简单的夹—仓嵌入式芯片组装，以芯片通道内染料的移动距离为定量检测信号，同时结合比色和热成像信号，实现了芯片上PSA免疫检测信号的肉眼可视三模式定量输出。在不依赖复杂分析仪器的情况下，该芯片对PSA的检测限（LOD）为1.4～2.8 ng/mL，低于实际前列腺癌的临床诊断阈值（4.0 ng/mL）。采用标准加样法，该芯片检测正常人血清中PSA浓度的加样回收率为90.5～105%。该项目为免疫检测纸芯片的研究提供了一种新思路，为光热效应在免疫检测纸芯片中的应用提供了理论基础和实验依据，在肿瘤标志物的现场即时检测中具有一定的应用潜力。

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