Beijing National Laboratory for Molecular Sciences, MOE Key Laboratory of Bioorganic Chemistry and Molecular Engineering, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Beijing National Laboratory for Molecular Sciences, MOE Key Laboratory of Bioorganic Chemistry and Molecular Engineering, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Biosens Bioelectron. 2014 May 15;55:438-45. doi: 10.1016/j.bios.2013.12.056. Epub 2014 Jan 3.
We demonstrate a novel fluorescent sensor for real-time and continuous monitoring of the variation of bisulfide in microdialysis effluents by using a nanoparticle-glutathione-fluorescein isothiocyanate (AuNP-GSH-FITC) probe coupled with on-line droplet-based microfluidic chip. The AuNP-GSH-FITC fluorescent probe was firstly developed and used for bisulfide detection in bulk solution by quantitative real-time PCR, which achieved a linear working range from 0.1 μM to 5.0 μM and a limit of detection of ~50 nM. The response time was less than 2 min. With the aid of co-immobilized thiol-polyethylene glycol, the probe exhibited excellent stability and reproducibility in high salinity solutions, including artificial cerebrospinal fluids (aCSF). By adding 0.1% glyoxal to the probe solution, the assay allowed quantification of bisulfide in the presence of cysteine at the micro-molarity level. Using the AuNP-GSH-FITC probe, a droplet-based microfluidic fluorescent sensor was further constructed for online monitoring of bisulfide variation in the effluent of microdialysis. By using fluorescence microscope-charge-coupled device camera as the detector, the integrated microdialysis/microfluidic chip device achieved a detection limit of 2.0 μM and a linear response from 5.0 μM to 50 μM for bisulfide in the tested sample. The method was successfully applied for the on-line measurement of bisulfide variation in aCSF and serum samples. It will be a very useful tool for tracking the variation of bisulfide or hydrogen sulfide in extracellular fluids.
我们展示了一种新颖的荧光传感器，用于通过使用纳米粒子-谷胱甘肽-荧光素异硫氰酸酯（AuNP-GSH-FITC）探针与在线液滴微流控芯片结合，实时连续监测微透析流出物中硫氢根的变化。首先开发了 AuNP-GSH-FITC 荧光探针，并通过定量实时 PCR 用于 bulk solution 中的硫氢根检测，其线性工作范围为 0.1 μM 至 5.0 μM，检测限约为 50 nM。响应时间小于 2 分钟。在共固定硫醇-聚乙二醇的帮助下，探针在包括人工脑脊液（aCSF）在内的高盐溶液中表现出出色的稳定性和重现性。通过向探针溶液中添加 0.1%乙二醛，可以在微摩尔级别的半胱氨酸存在下定量检测硫氢根。使用 AuNP-GSH-FITC 探针，进一步构建了基于液滴的微流荧光传感器，用于在线监测微透析流出物中硫氢根的变化。通过使用荧光显微镜-电荷耦合器件相机作为检测器，集成的微透析/微流控芯片装置实现了检测限为 2.0 μM，测试样品中硫氢根的线性响应范围从 5.0 μM 到 50 μM。该方法成功应用于 aCSF 和血清样品中硫氢根变化的在线测量。它将成为跟踪细胞外液中硫氢根或硫化氢变化的非常有用的工具。
