光图案化纳米孔薄膜集成微流控芯片的适配体亲和电泳研究  

Integrated Microfluidic Chip for Aptamer Affinity Probe ElectrophoresisAssay Using in Situ Photopatterned Nanoporous Membrane

作　　者：熊孟[1,3] 任金芝 胡善文 徐静娟 陈洪渊[3] XIONG Meng;REN Jin-Zhi;HU Shan-Wen;XU Jing-Juan;CHEN Hong-Yuan(School of Biotechnology,Jiangsu University of Science and Technology,Zhenjiang 212018,China;School of Public Health,Fujian Medical University,Fuzhou 350122,China;State Key Laboratory of Analytical Chemistry for Life Science,School of Chemistry and Chemical Engineering,Nanjing University,Nanjing 210023,China)

机构地区：[1]江苏科技大学生物技术学院,镇江212018 [2]福建医科大学公共卫生学院,福州350122 [3]南京大学生命分析化学国家重点实验室,南京大学化学化工学院,南京210023

出　　处：《分析化学》2021年第4期602-610,共9页Chinese Journal of Analytical Chemistry

基　　金：国家自然科学基金项目(Nos.21705059,21804064)资助。

摘　　要：设计了一种集成微流控芯片,基于适配体亲和毛细管电泳技术分析了痕量蛋白质。此集成芯片包含两个功能性单元,即用于样品富集的纳米孔薄膜单元和用于样品分离的凝胶筛分介质单元,前者可以提高检测灵敏度,后者可实现复合物有效分离。利用空间可控光图案化技术,在玻璃芯片的特定区域制备聚N-异丙基丙烯酰胺纳米孔薄膜微结构,并在分离通道与薄膜相邻处无缝灌注甲基纤维素凝胶,完成零死体积功能性单元集成。以凝血酶适配体亲和体系为模型,评估集成芯片的性能,以一种不含富集单元的简单双“T”微流控芯片为对照,采用共聚焦激光诱导荧光系统对其进行检测,经过30 s富集和120 s分离,集成芯片检测范围为1~40 nmol/L,检出限为1 nmol/L(S/N=3),比对照芯片的检出限(100 nmol/L)低100倍,显示出明显的信号增强能力和较好的分离度。此集成芯片有望用于其它亲和电泳体系,以提高生化分析检测灵敏度。Herein we proposed an integrated microfluidic chip to perform an aptamer-based affinity electrophoresis assay for the analysis of trace-level protein.The integrated platform included two functional units,a nanoporous membrane for sample preconcentration to improve detection sensitivity and a long gel sieving matrix for effective sample separation.The spatially controlled photopatterning technology was used to create poly(N-isopropylacrylamide)(PNIPAAm)nanoporous membrane microstructure inside the glass microchip,and methylcellulose(MC)gel solution was perfused adjacent to the membrane in the separation channel enabled zero dead volume.A model aptamer targeting thrombin was demonstrated to evaluate the performance of integrated microchip,and a simple double-T microchip was designed as a reference without the preconcentration unit.A confocal laser-induced fluorescence detection system(LIF)was utilized to quantitate the target protein thrombin.After 30 s of preconcentration and 120 s of separation,the integrated microfluidic chip showed significant signal enhancement and good separation resolution of thrombin complex with a detection range of 1-40 nmol/L and a detection limit of 1 nmol/L(S/N=3),which was over 100-fold lower than a simple double-T microchip(the detection limit was 100 nmol/L,S/N=3).The integrated platform presented in this work is anticipated to improve sensitivity of other affinity electrophoresis-based analytes.

关 键 词：光图案化 聚N-异丙基丙烯酰胺 预富集 集成微流控芯片 适配体亲和毛细管电泳 

分 类 号：TN492[电子电信—微电子学与固体电子学]