Fluorescence lifetime plate reader: Resolution and precision meet high-throughput

Karl J. Petersen, Kurt C. Peterson, Joseph M. Muretta, Sutton E. Higgins, Gregory D. Gillispie, David D. Thomas

Research output: Contribution to journalReview articlepeer-review

24 Scopus citations

Abstract

We describe a nanosecond time-resolved fluorescence spectrometer that acquires fluorescence decay waveforms from each well of a 384-well microplate in 3 min with signal-to-noise exceeding 400 using direct waveform recording. The instrument combines high-energy pulsed laser sources (5-10 kHz repetition rate) with a photomultiplier and high-speed digitizer (1 GHz) to record a fluorescence decay waveform after each pulse. Waveforms acquired from rhodamine or 5-((2-aminoethyl)amino) naphthalene-1-sulfonic acid dyes in a 384-well plate gave lifetime measurements 5- to 25-fold more precise than the simultaneous intensity measurements. Lifetimes as short as 0.04 ns were acquired by interleaving with an effective sample rate of 5 GHz. Lifetime measurements resolved mixtures of single-exponential dyes with better than 1% accuracy. The fluorescence lifetime plate reader enables multiple-well fluorescence lifetime measurements with an acquisition time of 0.5 s per well, suitable for high-throughput fluorescence lifetime screening applications.

Original languageEnglish (US)
Article number113101
JournalReview of Scientific Instruments
Volume85
Issue number11
DOIs
StatePublished - Nov 1 2014

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