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Fluorescence Lifetime Macro Imager for Biomedical Applications
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Published on: April 7, 2023

Long wavelength fluorescence lifetime standards for front-face fluorometry.

Bryan J McCranor1, Richard B Thompson

  • 1Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, 108 N. Greene Street, Baltimore, MD 21201-1503, USA.

Journal of Fluorescence
|December 3, 2009
PubMed
Summary

Developing new fluorescence lifetime standards is crucial for accurate measurements. This study presents a versatile Förster resonance energy transfer (FRET) method for creating reliable lifetime standards across various wavelengths, especially in the red and near-infrared spectrum.

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Area of Science:

  • Analytical Chemistry
  • Biophysics
  • Optical Engineering

Background:

  • Fluorescence lifetime measurements are increasingly used in various scientific instruments, including sensors and microscopy.
  • Accurate calibration standards are essential for the reliability of fluorescence lifetime-based devices.
  • Existing scattering-based standards are unsuitable for front-face excitation geometries, and a need exists for fluorescent lifetime standards, particularly in the red and near-infrared regions.

Purpose of the Study:

  • To develop a versatile method for creating fluorescent lifetime standards applicable to a wide range of wavelengths.
  • To address the limitations of current standards for front-face excitation geometries.
  • To provide reliable standards for long-wavelength fluorophores used in red and near-infrared spectroscopy.

Main Methods:

  • Utilized Förster resonance energy transfer (FRET) as the core mechanism for developing lifetime standards.
  • Designed standards with broad decreases in observed fluorescence lifetime.
  • Focused on achieving emission wavelengths in the red to near-infrared spectrum.

Main Results:

  • Successfully developed a FRET-based approach for creating fluorescence lifetime standards.
  • The developed standards exhibit brightness and high reproducibility.
  • The standards demonstrate a significant decrease in observed lifetime and emit in the red to near-infrared range.
  • The method is adaptable for creating standards in other wavelength regimes.

Conclusions:

  • The FRET-based approach provides a robust and adaptable method for generating essential fluorescence lifetime standards.
  • These new standards are suitable for various applications, including laboratory and field use, particularly with long-wavelength fluorophores.
  • This work addresses a critical need for reliable calibration tools in fluorescence spectroscopy and imaging.