Two-photon fluorescence imaging of intracellular hydrogen peroxide with chemoselective fluorescent probes Academic Article uri icon


MeSH Major

  • Fluorescent Dyes
  • Hydrogen Peroxide
  • Microscopy, Fluorescence
  • Molecular Imaging


  • We present the application of two-photon fluorescence (TPF) imaging to monitor intracellular hydrogen peroxide (H₂O₂) production in brain cells. For selective imaging of H₂O₂ over other reactive oxygen species, we employed small-molecule fluorescent probes that utilize a chemoselective boronate deprotection mechanism. Peroxyfluor-6 acetoxymethyl ester detects global cellular H₂O₂ and mitochondria peroxy yellow 1 detects mitochondrial H₂O₂. Two-photon absorption cross sections for these H₂O₂ probes are measured with a mode-locked Ti:sapphire laser in the wavelength range of 720 to 1040 nm. TPF imaging is demonstrated in the HT22 cell line to monitor both cytoplasmic H₂O₂ and localized H₂O₂ production in mitochondria. Endogenous cytoplasmic H₂O₂ production is detected with TPF imaging in rat astrocytes modified with d-amino acid oxidase. The TPF H₂O₂ imaging demonstrated that these chemoselective probes are powerful tools for the detection of intracellular H₂O₂.

publication date

  • November 6, 2013



  • Academic Article



  • eng

PubMed Central ID

  • PMC3787808

Digital Object Identifier (DOI)

  • 10.1117/1.JBO.18.10.106002

PubMed ID

  • 24084856

Additional Document Info

start page

  • 106002


  • 18


  • 10