Nortz Samuel P, Gupta Vanshika, Dick Jeffrey E
Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA.
Elmore Family School of Electrical and Computer Engineering, Purdue University, West Lafayette, IN 47906, USA.
Analyst. 2025 Apr 22;150(9):1795-1806. doi: 10.1039/d5an00017c.
Electrochemistry has become a key technique for studying biomolecular reactions and dynamics of living systems by using electron-transfer reactions to probe the complex interactions between biological redox molecules and their surrounding environments. To enable such measurements, redox mediators such as ferro/ferricyanide, ferrocene methanol, and tris(bipyridine) ruthenium(II) chloride are used. However, the impact of these exogeneous redox mediators on the health of cell cultures remains underexplored. Herein, we present the effects of three common redox mediators on the health of four of the most commonly used cell lines (Panc1, HeLa, U2OS, and MDA-MB-231) in biological studies. Cell health was assessed using three independent parameters: reactive oxygen species quantification by fluorescence flow cytometry, cell migration through scratch assays, and cell growth luminescence assays. We show that as the concentration of mediator exceeds 1 mM, ROS increases in all cell types while cell viability plumets. In contrast, cell migration was only hindered at the highest concentration of each mediator. Our observations highlight the crucial role that optimized mediator concentrations play in ensuring accuracy when studying biological systems by electrochemical methods. As such, these findings provide a critical reference for selecting redox mediator concentrations for bioanalytical studies on live cells.
电化学已成为通过利用电子转移反应来探测生物氧化还原分子与其周围环境之间复杂相互作用，从而研究生物分子反应和生命系统动力学的关键技术。为了实现此类测量，人们使用了诸如铁氰化亚铁/铁氰化铁、二茂铁甲醇和三（联吡啶）钌（II）氯化物等氧化还原介质。然而，这些外源性氧化还原介质对细胞培养健康的影响仍未得到充分研究。在此，我们展示了三种常见氧化还原介质对生物学研究中四种最常用细胞系（Panc1、HeLa、U2OS和MDA-MB-231）健康的影响。使用三个独立参数评估细胞健康状况：通过荧光流式细胞术进行活性氧物种定量、通过划痕试验进行细胞迁移以及通过发光试验进行细胞生长。我们发现，当介质浓度超过1 mM时，所有细胞类型中的活性氧都会增加，而细胞活力则会下降。相比之下，仅在每种介质的最高浓度下细胞迁移才会受到阻碍。我们的观察结果突出了优化介质浓度在通过电化学方法研究生物系统时确保准确性方面所起的关键作用。因此，这些发现为在活细胞生物分析研究中选择氧化还原介质浓度提供了重要参考。
