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The self-referencing oxygen-selective microelectrode: Detection of transmembrane oxygen flux from single cells

The self-referencing oxygen-selective microelectrode: Detection of transmembrane oxygen flux from single cells
The self-referencing oxygen-selective microelectrode: Detection of transmembrane oxygen flux from single cells
A self-referencing, polarographic, oxygen-selective microelectrode was developed for measuring oxygen fluxes from single cells. This technique is based on the translational movement of the microelectrode at a known frequency through an oxygen gradient, between known points, The differential current of the electrode was converted into a directional measurement of flux using the Fick equation. Operational characteristics of the technique were determined using artificial gradients. Calculated oxygen flux values matched theoretical values derived from static measurements. A test preparation, an isolated neuron, yielded an oxygen flux of 11.46+/-1.43 pmol cm(-2) s(-1) (mean +/- S.E.M.), a value in agreement with those available in the literature for single cells. Microinjection of metabolic substrates or a metabolic uncoupler increased oxygen flux, whereas microinjection of KCN decreased oxygen flux. In the filamentous alga Spirogyra greveilina, the probe could easily differentiate a 16.6 % difference in oxygen flux with respect to the position of the spiral chloroplast (13.3+/-0.4 pmol cm(-2) s(-1) at the chloroplast and 11.4+/-0.4 pmol cm(-2) s(-1) between chloroplasts), despite the fact that these positions averaged only 10.6+/-1.8 mu m apart (means +/- S.E.M.). A light response experiment showed realtime changes in measured oxygen flux correlated with changes in lighting. Taken together, these results show that the self-referencing oxygen microelectrode technique can be used to detect local oxygen fluxes with a high level of sensitivity and spatial resolution in real time. The oxygen fluxes detected reliably correlated with the metabolic state of the cell.
0022-0949
211-218
Land, S.C.
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Porterfield, D.M.
90790fa2-b4d8-4d76-94b6-cb50591c1c2a
Sanger, R.H.
ec3d4693-8abe-4cd0-89c0-02f44ebb056b
Smith, P.J.
003de469-9420-4f12-8f0e-8e8d76d28d6c
Land, S.C.
ab28350f-9bd5-4f11-a1c1-ee8164fa22b8
Porterfield, D.M.
90790fa2-b4d8-4d76-94b6-cb50591c1c2a
Sanger, R.H.
ec3d4693-8abe-4cd0-89c0-02f44ebb056b
Smith, P.J.
003de469-9420-4f12-8f0e-8e8d76d28d6c

Land, S.C., Porterfield, D.M., Sanger, R.H. and Smith, P.J. (1999) The self-referencing oxygen-selective microelectrode: Detection of transmembrane oxygen flux from single cells. Journal of Experimental Biology, 202 (2), 211-218. (PMID:9851909)

Record type: Article

Abstract

A self-referencing, polarographic, oxygen-selective microelectrode was developed for measuring oxygen fluxes from single cells. This technique is based on the translational movement of the microelectrode at a known frequency through an oxygen gradient, between known points, The differential current of the electrode was converted into a directional measurement of flux using the Fick equation. Operational characteristics of the technique were determined using artificial gradients. Calculated oxygen flux values matched theoretical values derived from static measurements. A test preparation, an isolated neuron, yielded an oxygen flux of 11.46+/-1.43 pmol cm(-2) s(-1) (mean +/- S.E.M.), a value in agreement with those available in the literature for single cells. Microinjection of metabolic substrates or a metabolic uncoupler increased oxygen flux, whereas microinjection of KCN decreased oxygen flux. In the filamentous alga Spirogyra greveilina, the probe could easily differentiate a 16.6 % difference in oxygen flux with respect to the position of the spiral chloroplast (13.3+/-0.4 pmol cm(-2) s(-1) at the chloroplast and 11.4+/-0.4 pmol cm(-2) s(-1) between chloroplasts), despite the fact that these positions averaged only 10.6+/-1.8 mu m apart (means +/- S.E.M.). A light response experiment showed realtime changes in measured oxygen flux correlated with changes in lighting. Taken together, these results show that the self-referencing oxygen microelectrode technique can be used to detect local oxygen fluxes with a high level of sensitivity and spatial resolution in real time. The oxygen fluxes detected reliably correlated with the metabolic state of the cell.

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Published date: January 1999
Organisations: University of Southampton

Identifiers

Local EPrints ID: 190525
URI: http://eprints.soton.ac.uk/id/eprint/190525
ISSN: 0022-0949
PURE UUID: 8d6e8408-3f5e-486e-8241-c9172e91b141
ORCID for P.J. Smith: ORCID iD orcid.org/0000-0003-4400-6853

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Date deposited: 13 Jun 2011 12:58
Last modified: 03 Dec 2019 01:40

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Contributors

Author: S.C. Land
Author: D.M. Porterfield
Author: R.H. Sanger
Author: P.J. Smith ORCID iD

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