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Anal Biochem. 2002 Oct 1;309(1):19-26.

Real-time background suppression during frequency domain lifetime measurements.

Author information

  • 1Center for Fluorescence Spectroscopy, University of Maryland at Baltimore, Department of Biochemistry and Molecular Biology, 725 West Lombard Street, Baltimore, MD 21201, USA.

Erratum in

  • Anal Biochem. 2002 Dec 1;311(1):102. Maliwal, Baldri P [corrected to Maliwal, Badri P].


We describe real time background suppression of autofluorescence from biological samples during frequency domain or phase modulation measurements of intensity decays. For these measurements the samples were excited with a train of light pulses with widths below 1 ps. The detector was gated off for a short time period of 10 to 40 ns during and shortly after the excitation pulse. The reference signal needed for the frequency domain measurement was provided by a long-lifetime reference fluorophore which continues to emit following the off-gating pulse. Both the sample and the reference were measured under identical optical and electronic conditions avoiding the need for correction of the photomultiplier tube signal for the gating sequence. We demonstrate frequency domain background suppression using a mixture of short- and long-lifetime probes and for a long-lifetime probe in human plasma with significant autofluorescence.

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