Heterodyning of modulated pulses for fluorescence lifetime measurements in flow cytometry

Bertram G. Pinsky; John J. Ladasky

doi:10.1117/12.182706

17 August 1994 Heterodyning of modulated pulses for fluorescence lifetime measurements in flow cytometry

Bertram G. Pinsky, John J. Ladasky

Proceedings Volume 2137, Time-Resolved Laser Spectroscopy in Biochemistry IV; (1994) https://doi.org/10.1117/12.182706
Event: OE/LASE '94, 1994, Los Angeles, CA, United States

Abstract

Heterodyning of modulated pulses in flow cytometry in the PMT offers three major advantages over external homodyning techniques: first, intermodulation distortion is reduced; second, the ability to choose a lower intermediate frequency for evaluating phase affords the opportunity to use digital signal processing techniques to measure phase; third, the limiters we used previously in homodyning phase measurement circuitry may be removed, thereby reducing phase distortion. We constructed a heterodyning PMT base capable of providing an intermediate frequency of 1 MHz, and installed it on our phase flow cytometer (PFC). The fluorescence lifetime of 2 micrometers Fluoresbrite yellow beads was approximately 3 to 4 ns on the heterodyning system, which compares well to the 3.5 ns measured with the original homodyning PFC electronics. This lifetime parameter, when coupled with recently-developed lifetime-based probes for Ca²⁺ and pH, provides new opportunities in flow cytometry for study of intensity-independent intracellular physiology.

Citation Download Citation

Bertram G. Pinsky and John J. Ladasky "Heterodyning of modulated pulses for fluorescence lifetime measurements in flow cytometry", Proc. SPIE 2137, Time-Resolved Laser Spectroscopy in Biochemistry IV, (17 August 1994); https://doi.org/10.1117/12.182706

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