Visualization of intracellular hydrogen peroxide with HyPer, a genetically encoded fluorescent probe

Nataliya M Mishina; Kseniya N Markvicheva; Dmitry S Bilan; Mikhail E Matlashov; Marina V Shirmanova; David Liebl; Carsten Schultz; Sergey Lukyanov; Vsevolod V Belousov

doi:10.1016/B978-0-12-405883-5.00003-X

Visualization of intracellular hydrogen peroxide with HyPer, a genetically encoded fluorescent probe

Methods Enzymol. 2013:526:45-59. doi: 10.1016/B978-0-12-405883-5.00003-X.

Authors

Nataliya M Mishina¹, Kseniya N Markvicheva, Dmitry S Bilan, Mikhail E Matlashov, Marina V Shirmanova, David Liebl, Carsten Schultz, Sergey Lukyanov, Vsevolod V Belousov

Affiliation

¹ Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, RAS, Moscow, Russia.

PMID: 23791093
DOI: 10.1016/B978-0-12-405883-5.00003-X

Abstract

The fluorescent sensor HyPer allows monitoring of intracellular H2O2 levels with a high degree of sensitivity and specificity. Here, we provide a detailed protocol of ratiometric imaging of H2O2 produced by cells during phagocytosis, including instructions for experiments on different commercial confocal systems, namely, Leica SP2, Leica SP5, and Carl Zeiss LSM, as well as wide-field Leica 6000 microscope. The general experimental scheme is easily adaptable for imaging H2O2 production by various cell types under a variety of conditions.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Cell Line
Fluorescent Dyes / metabolism*
Humans
Hydrogen Peroxide / analysis*
Hydrogen Peroxide / metabolism
Microscopy, Confocal / methods*
Optical Imaging / methods
Phagocytosis
Transfection / methods

Substances

Fluorescent Dyes
Hydrogen Peroxide