AAT Bioquest

What is the difference between ethidium homodimer and dihydroethidium in terms of staining?

Posted April 24, 2020


Ethidium homodimer is generally used as a DNA stain for dead cells. Due the positive charge of ethidium bromide, it has low membrane permeability and can only penetrate a cell membrane after it is damaged. Once inside the cell, the dye then fluoresces upon binding to DNA.


Dihydroethidium, which is a reduced form of ethidium bromide, is commonly used an indicator for ROS (reactive oxygen species), mainly superoxide. Once within the cell, dihydroethidium, which initially has a blue color, fluoresces red when oxidized. Due to the dye’s affinity for DNA, the nucleus is stained red.

Additional resources

Hydroethidine [Dihydroethidium] *CAS 104821-25-2*

Edwards, J.R., Diamantakos, E.A., Peuler, J.D., Lamar, P.C., & Prozialeck, W.C. (2007). A novel method for the evaluation of proximal tubule epithelial cellular necrosis in the intact rat kidney using ethidium homodimer. BMC Physiology, 7, 1. doi:10.1186/1472-6793-7-1

Peshavariya H.M., Dusting, G.J., & Selemidis, S. (2007). Analysis of dihydroethidium fluorescence for the detection of intracellular and extracellular superoxide produced by NADPH oxidase. Free Radical Research, 41(6), 699-712. doi:10.1080/10715760701297354