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bioRxiv · 10.1101/2023.04.24.538172

Can Single Cell Respiration be Measured by Scanning Electrochemical Microscopy (SECM)?

Cremin, K.·Meloni, G.·Valavanis, D.·soyer, o. s.·Unwin, P. R.
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Abstract

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) at 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) potentials 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 1x10-18 to 1x10-16 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. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/538172v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@14a6e59org.highwire.dtl.DTLVardef@192d44borg.highwire.dtl.DTLVardef@afb2caorg.highwire.dtl.DTLVardef@103131b_HPS_FORMAT_FIGEXP M_FIG C_FIG

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Cremin, K., Meloni, G., Valavanis, D., soyer, o. s., Unwin, P. R.. 2023-04-26. Can Single Cell Respiration be Measured by Scanning Electrochemical Microscopy (SECM)?. https://doi.org/10.1101/2023.04.24.538172

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