Department of Mechanical and Aerospace Engineering, Monash University, Room 227, New Horizons Building, 20 Research Way, Clayton, Melbourne, Victoria 3800, Australia.
Monash Institute of Pharmaceutical Sciences (MIPS), Monash University, 381 Royal Parade, Parkville, Melbourne, Victoria 3052, Australia.
Anal Chem. 2022 Jan 25;94(3):1726-1732. doi: 10.1021/acs.analchem.1c04312. Epub 2022 Jan 11.
It is well known that hydrogen peroxide (HO) is a signaling molecule essential for vital physiological reactions in mammalian cells, such as cell survival, intercellular communication, and cancer metabolism. However, to fully understand the function of HO, it is critical to monitor its intracellular and/or extracellular concentrations. Current techniques implemented to address this need require large sample volumes, expensive instrumentation, and long sample preparation and analysis times, inapplicable to inline or online monitoring. In this paper, a new integrated microfluidic device capable of overcoming these limitations is demonstrated for the colorimetric detection of extracellular hydrogen peroxide HO. The device contains an optical waveguide to determine absorbance changes and micromixers to enable complete mixing of reagents using a passive approach. This novel HO-sensing device has allowed the detection of HO in the range of 0.5-60 μM with a detection limit of 167 ± 5.8 nM and a sensitivity of 13.5 ± 0.1 AU/mM. Proof of concept of the device was demonstrated by quantifying HO release from benign prostatic epithelial (BPH-1) cells upon stimulation with phorbol 12-myristate 13-acetate (PMA). Results show that this integrated device can be potentially utilized to continuously monitor cell-released metabolites autonomously without constant human supervision during the process. Furthermore, this can be achieved without interfering with the cell culture conditions, as only a very small volume of conditioned media (less than 0.4 μL), and not the cells, is required.
众所周知，过氧化氢（HO）是哺乳动物细胞中重要的信号分子，对于细胞存活、细胞间通讯和癌症代谢等重要的生理反应必不可少。然而，要全面了解 HO 的功能，关键是要监测其细胞内和/或细胞外浓度。目前用于满足这一需求的技术需要大量的样本量、昂贵的仪器设备以及较长的样本准备和分析时间，无法用于在线或实时监测。本文展示了一种新的集成微流控装置，该装置能够克服这些限制，用于检测细胞外过氧化氢（HO）的比色法检测。该装置包含一个光学波导，用于确定吸光度变化，以及微混合器，用于通过被动方法实现试剂的完全混合。这种新型的 HO 传感装置能够检测 0.5-60 μM 范围内的 HO，检测限为 167 ± 5.8 nM，灵敏度为 13.5 ± 0.1 AU/mM。通过用佛波醇 12-肉豆蔻酸 13-乙酸酯（PMA）刺激良性前列腺上皮（BPH-1）细胞来验证该装置的概念验证。结果表明，该集成装置可潜在用于在无人持续监督的情况下自主连续监测细胞释放的代谢物，而无需对细胞培养条件造成干扰，因为仅需要非常少量的条件培养基（小于 0.4 μL），而不是细胞。
