扫描电化学显微镜(SECM)能测量单细胞呼吸吗?

IF 4.6 Q1 CHEMISTRY, ANALYTICAL ACS Measurement Science Au Pub Date : 2023-07-10 DOI:10.1021/acsmeasuresciau.3c00019
Kelsey Cremin, Gabriel N. Meloni, Dimitrios Valavanis, Orkun S. Soyer* and Patrick R. Unwin*, 
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摘要

超微电极(UME)或等效微电极探针越来越多地用于单细胞测量细胞特性和过程,包括生理活动,如代谢通量和呼吸率。这种测量灵敏度的主要挑战包括:(i)细胞和UME通量的相对大小(表现在电流中);以及(ii)UME响应随时间变化的稳定性问题。为了探索这些因素对电化学细胞测量精度的影响程度,我们对测量条件和实验参数进行了系统分析,以通过单HeLa细胞中的耗氧率(OCR)确定单细胞呼吸率。使用以铂UME为探针的扫描电化学显微镜(SECM),我们采用了一种自参考测量协议,该协议在SECM中很少使用,通过该协议,UME从本体溶液到电池重复接近,并在电池附近和本体溶液中进行短脉冲氧还原反应(ORR)电位。这种方法能够周期性地跟踪近电池响应与之重复比较(参考)的批量UME响应,并且还确保仅短暂地执行电池附近的ORR,从而最大限度地减少电化学过程对电池的影响。SECM实验与有限元法(FEM)建模框架相结合,以模拟氧气扩散和UME响应。将单细胞OCR的实际范围设为1×10-18至1×10-16mol s-1,FEM模拟和自参考SECM测量相结合的结果表明,这些OCR值等于或低于该技术目前的检测灵敏度。我们为未来改进这些测量提供了一组基于模型的建议,但强调如果要实现单细胞OCR测量,就需要在SECM测量的稳定性和精度方面进行非凡的改进。
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Can Single Cell Respiration be Measured by Scanning Electrochemical Microscopy (SECM)?

Ultramicroelectrode (UME), or, equivalently, microelectrode, probes are increasingly used for single-cell measurements of cellular properties and processes, including physiological activity, such as metabolic fluxes and respiration rates. Major challenges for the sensitivity of such measurements include: (i) the relative magnitude of cellular and UME fluxes (manifested in the current); and (ii) issues around the stability of the UME response over time. To explore the extent to which these factors impact the precision of electrochemical cellular measurements, we undertake a systematic analysis of measurement conditions and experimental parameters for determining single cell respiration rates via the oxygen consumption rate (OCR) in single HeLa cells. Using scanning electrochemical microscopy (SECM), with a platinum UME as the probe, we employ a self-referencing measurement protocol, rarely employed in SECM, whereby the UME is repeatedly approached from bulk solution to a cell, and a short pulse to oxygen reduction reaction (ORR) potential is performed near the cell and in bulk solution. This approach enables the periodic tracking of the bulk UME response to which the near-cell response is repeatedly compared (referenced) and also ensures that the ORR near the cell is performed only briefly, minimizing the effect of the electrochemical process on the cell. SECM experiments are combined with a finite element method (FEM) modeling framework to simulate oxygen diffusion and the UME response. Taking a realistic range of single cell OCR to be 1 × 10–18 to 1 × 10–16 mol s–1, results from the combination of FEM simulations and self-referencing SECM measurements show that these OCR values are at, or below, the present detection sensitivity of the technique. We provide a set of model-based suggestions for improving these measurements in the future but highlight that extraordinary improvements in the stability and precision of SECM measurements will be required if single cell OCR measurements are to be realized.

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ACS Measurement Science Au
ACS Measurement Science Au 化学计量学-
CiteScore
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期刊介绍: ACS Measurement Science Au is an open access journal that publishes experimental computational or theoretical research in all areas of chemical measurement science. Short letters comprehensive articles reviews and perspectives are welcome on topics that report on any phase of analytical operations including sampling measurement and data analysis. This includes:Chemical Reactions and SelectivityChemometrics and Data ProcessingElectrochemistryElemental and Molecular CharacterizationImagingInstrumentationMass SpectrometryMicroscale and Nanoscale systemsOmics (Genomics Proteomics Metabonomics Metabolomics and Bioinformatics)Sensors and Sensing (Biosensors Chemical Sensors Gas Sensors Intracellular Sensors Single-Molecule Sensors Cell Chips Arrays Microfluidic Devices)SeparationsSpectroscopySurface analysisPapers dealing with established methods need to offer a significantly improved original application of the method.
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