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Second-Harmonic Generation Imaging of Membrane Potential with Photon Counting

Published online by Cambridge University Press:  06 November 2008

Jiang Jiang  and
Rafael Yuste
Jiang Jiang*
Affiliation:
HHMI, Department of Biological Sciences, Columbia University, New York, NY 10027, USA
Rafael Yuste
Affiliation:
HHMI, Department of Biological Sciences, Columbia University, New York, NY 10027, USA
*
Corresponding author. E-mail: jjiang@ibn.a-star.edu.sg
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Abstract

Second-harmonic generation (SHG) can be used for imaging membrane potential in neurons, but poor signal-to-noise (S/N) limits accurate measurements of small voltage transients. We use photon counting to improve the S/N of weak SHG signal detection. Photon counting generates shot-noise limited and integrable signals, eliminates pulse-to-pulse variation, and built-in discriminators reduces the background to practically zero. In single trials, by using photon counting, we obtain a more than a twofold S/N increase over analog voltage detection. Trial-to-trial variability is also reduced by 50%. Finally, we show that, using photon counting, the kinetics of fast events such as action potentials can be recorded more accurately.

Keywords

second-harmonic generationphoton countingneuronmembrane potential
Type
Multiphoton Microscopy–Special Section
Information
Microscopy and Microanalysis , Volume 14 , Issue 6 , December 2008 , pp. 526 - 531
Copyright
Copyright © Microscopy Society of America 2008

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References

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