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

Extended cavity laser enhanced two-photon flow cytometry.

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  • 1University of Michigan, Center for Ultrafast Optical Science, Electrical Engineering & Computer Science Department, Ann Arbor, Michigan 48109-2099, USA. etkaczyk@umich.edu

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.

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