Detecting oxygen consumption in the proximity of Saccharomyces cerevisiae cells using self‐assembled fluorescent nanosensors

Abstract

We describe a strategy for the preparation and self-assembly of fluorescent nanosensors onto Saccharomyces cerevisiae cell surfaces for dynamically measuring oxygen concentration in the proximity of living cells. Amine functionalized polystyrene nanobeads were impregnated with an oxygen-sensitive ruthenium(II) complex and the beads' surface was coated with polyethylenimine. The resulting nanosensors were assembled on individual S. cerevisiae cells in a controlled manner at physiological pH for continuously monitoring oxygen consumption. This approach exemplifies a general scheme for assembling fluorescent nanosensors on cells for the non-invasive, reversible, and real-time measurement of other physiologically relevant processes, such as the efflux of protons and carbon dioxide, or the influx of glucose.

A microwell array device capable of measuring single-cell oxygen consumption rates

“…Another technique for measuring the oxygen consumption in the near proximity of individual yeast cells coated with fluorescent oxygen nanobeads has also been presented. [17] While the method is relatively high throughput (~100 cells in parallel), it still only measures oxygen flux. In addition, the beads may interfere with normal cell behavior, and certain cell types such as macrophages can consume the beads, as we have found in our own studies.…”

Direct measurement of oxygen consumption rates from attached and unattached cells in a reversibly sealed, diffusionally isolated sample chamber

“…However, the drawbacks of electrodes include low sensitivities, signal drift, and consumption of oxygen by the electrode itself [10,11]. Other recent methods have entailed scanning electrochemical microscopy (SECM) and utilization of nanobead sensors attached to the outer membrane of cells [12,13]. However, these techniques have been employed in openly diffusible environments and only indicate oxygen flux near the cell membrane or the oxygen sensor.…”

Light-directed, spatially addressable oxygen detection in a hydrogel microarray based on phase-based lifetime detection using a digital micromirror device

“…However, these sensors are usually susceptible to membrane fouling, significant drift, analyte depletion and time-consuming because one electrode can only monitor one sample at a time. Alternatively, luminescent optical sensing, which is based on the ability of molecular oxygen to dynamically quench oxygen-sensitive photoluminescent dyes, offers a fast, linear response, and immunity to drift from the consumption of O 2 [5][6][7][8][9][10][11][12][13][14][15][16].…”

“…Quantification of oxygen concentrations with oxygen-sensitive photoluminescent dyes is measured by either the degree of luminescent intensity quenching [5][6][7][8][9] or the luminescent lifetime [10][11][12][13][14][15][16]. Detecting the luminescent lifetime to quantify oxygen concentrations has been proven to have a higher sensitivity due to the inherent stability of the signal.…”