Fluorescence lifetime standards for time and frequency domain fluorescence spectroscopy

Area of Science:

• Photochemistry and Photophysics
• Analytical Chemistry
• Spectroscopy

Background:

• Accurate fluorescence lifetime measurements are critical for various scientific disciplines.
• Establishing reliable fluorescence lifetime standards is essential for instrument calibration and validation.
• Existing standards may have limitations in terms of availability, photostability, or spectral range.

Purpose of the Study:

• To identify and validate a series of commercially available fluorophores as reliable fluorescence lifetime standards.
• To assess the suitability of these standards for both time-domain and frequency-domain fluorescence measurements.
• To provide a comprehensive dataset for instrument calibration and inter-laboratory comparison.

Main Methods:

• Multiple laboratories independently measured fluorescence decays of selected dyes in solution at 20°C.
• Utilized single-photon timing and multifrequency phase and modulation fluorometry techniques with laser excitation.
• Employed spectroscopic-grade solvents including methanol, cyclohexane, and water.

Main Results:

• A list of 14 photostable, soluble fluorophores suitable as fluorescence lifetime standards was compiled.
• Measured fluorescence lifetimes range from 89 picoseconds to 31.2 nanoseconds, covering a broad spectral range.
• Decay times were found to be independent of excitation and emission wavelengths, with good inter-laboratory agreement.

Conclusions:

• The identified fluorophores serve as robust standards for calibrating and testing time- and frequency-domain fluorescence instrumentation.
• Excellent correlation was observed between time-domain and frequency-domain lifetime measurements.
• These standards are valuable for accurate fluorescence lifetime determination and eliminating instrument-specific artifacts.