Extended cavity laser enhanced two-photon flow cytometry

J Biomed Opt. 2008 Jul-Aug;13(4):041319. doi: 10.1117/1.2967983.

Abstract

We demonstrate enhanced sensitivity in two-photon flow cytometry with an extended cavity laser excitation source. At low power, the home-built 20-MHz oscillator was able to detect a significantly larger fraction, in either phosphate buffered saline (PBS) or whole blood, of green fluorescent protein (GFP)-expressing MCA-207 cells cross-labeled with the membrane-binding lipophilic dye DiD. A geometrical model is used to explain unique features of the signals resulting from the different spatial distribution of DiD and GFP. These unique features include sub-square law scaling of unsaturated two-photon signal, a sigmoidal sensitivity curve for detection under varying powers for cell detection thresholds as low as a single photon, and uncorrelated signal strengths in two detection channels.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Equipment Design
  • Equipment Failure Analysis
  • Flow Cytometry / instrumentation*
  • Flow Cytometry / methods
  • Lasers*
  • Microscopy, Fluorescence, Multiphoton / instrumentation*
  • Microscopy, Fluorescence, Multiphoton / methods
  • Photometry / instrumentation*
  • Photons
  • Radiation Dosage
  • Radiometry / instrumentation*