Nat Methods. 2006 Apr;3(4):281-6.

Genetically encoded fluorescent indicator for intracellular hydrogen peroxide.

Belousov VV, Fradkov AF, Lukyanov KA, Staroverov DB, Shakhbazov KS, Terskikh AV, Lukyanov S.

Source

Laboratory of Genes for Regeneration, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya Street 16/10, Moscow 117997, Russia.

Abstract

We developed a genetically encoded, highly specific fluorescent probe for detecting hydrogen peroxide (H(2)O(2)) inside living cells. This probe, named HyPer, consists of circularly permuted yellow fluorescent protein (cpYFP) inserted into the regulatory domain of the prokaryotic H(2)O(2)-sensing protein, OxyR. Using HyPer we monitored H(2)O(2) production at the single-cell level in the cytoplasm and mitochondria of HeLa cells treated with Apo2L/TRAIL. We found that an increase in H(2)O(2) occurs in the cytoplasm in parallel with a drop in the mitochondrial transmembrane potential (DeltaPsi) and a change in cell shape. We also observed local bursts in mitochondrial H(2)O(2) production during DeltaPsi oscillations in apoptotic HeLa cells. Moreover, sensitivity of the probe was sufficient to observe H(2)O(2) increase upon physiological stimulation. Using HyPer we detected temporal increase in H(2)O(2) in the cytoplasm of PC-12 cells stimulated with nerve growth factor.

PMID:: 16554833; [PubMed - indexed for MEDLINE]

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