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Differential Imaging of Biological Structures with Doubly-resonant Coherent Anti-stokes Raman Scattering (CARS)
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Differential polarization nonlinear optical microscopy with adaptive optics controlled multiplexed beams.

Masood Samim1, Daaf Sandkuijl, Ian Tretyakov

  • 1Department of Physics and Institute for Optical Sciences, University of Toronto, 60 St. George Street, Toronto, ON M5S 1A7, Canada. virgis.barzda@utoronto.ca.

International Journal of Molecular Sciences
|September 12, 2013
PubMed
Summary

Differential polarization microscopy offers advanced structural analysis of biological materials. This technique precisely measures nonlinear optical signal anisotropy for detailed fiber orientation and molecular studies in dynamic samples.

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Area of Science:

  • Nonlinear optics
  • Microscopy
  • Biophysics

Background:

  • Ordered biological assemblies and microcrystalline aggregates require advanced structural investigation methods.
  • Nonlinear optical signal anisotropy measurements offer sensitive probing of microscopic organization.

Purpose of the Study:

  • To develop and present a novel differential polarization microscope for enhanced structural analysis.
  • To demonstrate the technique's application in studying collagen and cellulose structures.

Main Methods:

  • Utilizing time-multiplexed pulsed laser beams with perpendicular polarization and photon-counting detection.
  • Employing deformable membrane mirrors and genetic algorithms for aberration correction and beam overlap optimization.
  • Performing pixel-by-pixel polarization-resolved measurements with sub-diffraction-limited accuracy.

Main Results:

  • Successfully applied differential polarization microscopy to structural studies of collagen and cellulose.
  • Determined fiber orientation and structural properties in biological tissues.
  • Precisely determined the orientation of fluorescent molecules (Congo Red) labeling the fibers.

Conclusions:

  • Differential polarization microscopy is a powerful tool for structural investigations of ordered biological materials.
  • The technique overcomes limitations like photobleaching and sample movement, enabling dynamic in vivo studies.
  • Fast data acquisition allows real-time monitoring of nonlinear signal anisotropy changes in live biological structures.