Analysis of fluorescence lifetime data for single rhodamine molecules in flowing sample streams

Fluorescence events for individual molecules of Rhodamine 110 dye in a stream of methanol occur as bursts of 5-300 photoelectrons when the dye molecules transit the volume interrogated by the pulsed excitation laser. The arrival times for these events, extracted by use of time-correlated single-photon counting techniques, are distributed exponentially over a ∼10-ns time window gated to discriminate against prompt scattered light. Still, residual scattered light and solvent-impurity fluorescence contribute significantly to the recorded time distributions, which therefore must be analyzed as sums of an exponential decay and a background. Results obtained from two-parameter nonlinear least-squares fits are compared with maximum-likelihood estimates which ignore the background

[1]  W. P. Ambrose,et al.  Fluorescence spectroscopy and spectral diffusion of single impurity molecules in a crystal , 1991, Nature.

[2]  M Köllner,et al.  How to find the sensitivity limit for DNA sequencing based on laser-induced fluorescence. , 1993, Applied optics.

[3]  J. Tellinghuisen.  Effect of background on the least-squares estimation of exponential decay parameters , 1993 .

[4]  Steven A. Soper,et al.  Detection and identification of single molecules in solution , 1992 .

[5]  Peter Hall,et al.  Better estimates of exponential decay parameters , 1981 .

[6]  D. A. Lewis,et al.  High resolution laser spectroscopy with minute samples , 1977 .

[7]  Jürgen Wolfrum,et al.  How many photons are necessary for fluorescence-lifetime measurements? , 1992 .

[8]  Better Estimates of Multiexponential Decay Parameters , 1984 .

[9]  R. Keller,et al.  Single-molecule detection as an approach to rapid DNA sequencing. , 1992, Trends in biotechnology.

[10]  W. Fairbank,et al.  Detection of individual atoms in helium buffer gas and observation of their real-time motion. , 1980, Optics letters.

[11]  Richard A. Keller,et al.  Detection and lifetime measurement of single molecules in flowing sample streams by laser‐induced fluorescence , 1993 .

[12]  Quincy L. Mattingly,et al.  Photon Burst Detection of Single Near-Infrared Fluorescent Molecules , 1993 .

[13]  Steven A. Soper,et al.  Laser-Induced Fluorescence Detection of Rhodamine-6G at 6 × 10−15 M , 1991 .

[14]  Charles W. Wilkerson,et al.  Bias and precision in the estimation of exponential decay parameters from sparse data , 1993 .

[15]  J. Longworth,et al.  Late Output Pulses from Fast Photomultipliers , 1972 .

[16]  James H. Jett,et al.  SINGLE-MOLECULE DETECTION OF RHODAMINE 6G IN ETHANOLIC SOLUTIONS USING CONTINUOUS WAVE LASER EXCITATION , 1991 .

[17]  Steven A. Soper,et al.  Detection of single fluorescent molecules , 1990 .

[18]  W. Moerner,et al.  Optical detection and spectroscopy of single molecules in a solid. , 1989, Physical review letters.

[19]  J. Jett,et al.  High-speed DNA sequencing: an approach based upon fluorescence detection of single molecules. , 1989, Journal of biomolecular structure & dynamics.

[20]  Alfons Penzkofer,et al.  Fluorescence quenching of rhodamine 6G in methanol at high concentration , 1986 .