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Detecting subtle plasma membrane perturbation in living cells using second harmonic generation imaging.

Erick K Moen1, Bennett L Ibey2, Hope T Beier2

  • 1Department of Electrical Engineering - Electrophysics, University of Southern California at Los Angeles, Los Angeles, California.

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|May 24, 2014
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Summary

Second harmonic generation (SHG) imaging visualizes cell membrane changes. This study shows SHG can detect rapid plasma membrane disruptions caused by electric fields, highlighting its potential for live cell analysis.

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Area of Science:

  • Biophysics
  • Cell Biology
  • Nonlinear Optics

Background:

  • Second harmonic generation (SHG) signals require broken centrosymmetry, making them suitable for studying interfacial layers like the cell plasma membrane.
  • Lipophilic SHG probes offer a way to visualize subtle changes in membrane structure.
  • Understanding plasma membrane dynamics is crucial for cell function and response to external stimuli.

Purpose of the Study:

  • To evaluate lipophilic SHG probes for detecting minor plasma membrane perturbations.
  • To assess the capability of SHG imaging to capture rapid changes in membrane symmetry.
  • To investigate the effects of nanosecond-pulsed electric fields on plasma membrane integrity using SHG.

Main Methods:

  • Screening of three lipophilic probes (Di-4-ANEPPDHQ, FM4-64, all-trans-retinol) for SHG effectiveness in Jurkat cells.
  • Application of nanosecond-pulsed electric fields to cells labeled with SHG probes.
  • Simultaneous acquisition of SHG and fluorescence signals to differentiate probe behavior.
  • Analysis of SHG signal changes in response to electric field-induced membrane perturbations.

Main Results:

  • Di-4-ANEPPDHQ demonstrated superior SHG signal strength and minimal photobleaching compared to other probes.
  • Exposure to nanosecond-pulsed electric fields caused an immediate ~50% reduction in SHG signal at the cell's anodic pole.
  • Signal changes were independent of probe diffusion, membrane potential, or fluidity, as indicated by fluorescence data.
  • The observed decrease in SHG signal is hypothesized to result from the disruption of the membrane's interfacial symmetry.

Conclusions:

  • SHG imaging, particularly with Di-4, is effective for visualizing plasma membrane dynamics.
  • The technique can detect rapid, subtle disturbances in plasma membrane structure induced by external factors like electric fields.
  • SHG imaging holds significant promise as a tool for real-time monitoring of plasma membrane integrity in living cells.