2 May 1997 Phase-sensitive detection as a means to recover fluoresence signals from interfering backgrounds in analytical cytology measurements
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Proceedings Volume 2982, Optical Diagnostics of Biological Fluids and Advanced Techniques in Analytical Cytology; (1997); doi: 10.1117/12.273642
Event: BiOS '97, Part of Photonics West, 1997, San Jose, CA, United States
Abstract
A phase-sensitive flow cytometer has ben developed to measure fluorescence emission signals from particles and cells labeled with fluorophores based on excited-state lifetimes and to quantify fluorescence lifetimes directly as a parameter in real time. This instrument combines flow cytometry (FCM) and frequency-domain, fluorescence lifetime spectroscopy measurement principles to provide unique features for making analytical cytology measurements in flow, while maintaining conventional FCM measurement capabilities. In this study we present the underlying theory of the phase-sensitive detector, along with the principles of phase-sensitive detection with respect to resolving fluorescence emission signal from background interferences caused by (1) Rayleigh scatter, (2) unbound/free dye, and (3) low-level intrinsic cellular autofluorescence, all based on differences in phase-shifts of the signals input to a phase-sensitive detector with respect to a fixed reference signal. Investigative studies directed at detecting and eliminating particle- and cell-associated and steady-state background interferences in fluorescence measurements are underway. Examples to illustrate the application of this new technology to biological problems encountered in flow cytometry applications will be presented.
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John A. Steinkamp, Bruce E. Lehnert, Jan F. Keij, "Phase-sensitive detection as a means to recover fluoresence signals from interfering backgrounds in analytical cytology measurements", Proc. SPIE 2982, Optical Diagnostics of Biological Fluids and Advanced Techniques in Analytical Cytology, (2 May 1997); doi: 10.1117/12.273642; http://dx.doi.org/10.1117/12.273642
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KEYWORDS
Luminescence

Signal detection

Phase shifts

Light scattering

Sensors

Electronic filtering

Modulation

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