Graphical representation and multicomponent analysis of single-frequency fluorescence lifetime imaging microscopy data

J Microsc. 2004 Jan;213(1):1-5. doi: 10.1111/j.1365-2818.2004.01265.x.

Abstract

Graphical representation of fluorescence lifetime imaging microscopy data demonstrates that a mixture of two components with single exponential decays can be resolved by single frequency measurements. We derive a method based on linear fitting that allows the calculation of the fluorescence lifetimes of the two components. We show that introduction of proper error-weighting results in a non-linear method that is mathematically identical to a global analysis algorithm that was recently derived. The graphical approach was applied to cellular data obtained from a lifetime-based phosphorylation assay for the epidermal growth factor receptor and yielded results similar to those obtained by a global analysis algorithm.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carbocyanines / metabolism*
  • Cell Line
  • ErbB Receptors / metabolism*
  • Fluorescence Resonance Energy Transfer
  • Fluorescent Dyes / metabolism*
  • Green Fluorescent Proteins
  • Humans
  • Image Processing, Computer-Assisted / methods*
  • Luminescent Proteins / metabolism*
  • Microscopy, Fluorescence / methods*
  • Phosphorylation
  • Phosphotyrosine / metabolism
  • Recombinant Fusion Proteins / metabolism
  • Tumor Cells, Cultured

Substances

  • Carbocyanines
  • Fluorescent Dyes
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • cyanine dye 3
  • Green Fluorescent Proteins
  • Phosphotyrosine
  • ErbB Receptors