AAT Bioquest

How does propidium iodide staining work?

Posted July 22, 2020


Propidium iodide (PI) is a cell-impermeant DNA binding dye that can be used to stain cells and nucleic acids. PI binds to DNA by intercalating between the bases with a stoichiometry of one dye per 4-5 base pairs of DNA. Little or no sequence preference is observed. Free PI has excitation/emission maximums of 493/636 nm, respectively. Once bound to DNA, the excitation/emission maximum is shifted to 535/617 nm, and its quantum yield is enhanced 20-30 fold, exhibiting bright orange-red fluorescence.

As a membrane impermeant dye, PI is generally excluded from viable cells. However, it can penetrate the dead cells, who has a compromised membrane integrity, and stain the intracellular double-stranded DNA molecules. Therefore, PI is widely used to evaluate cell viability, measure DNA content in cell cycle analysis, as well as visualize the nucleus and other DNA-containing organelles.

Additional resources

Propidium iodide *CAS 25535-16-4*

Live or Dead™ Cell Viability Assay Kit *Green/Red Dual Fluorescence*

Cell Meter™ Cell Viability Assay Kit *Blue Fluorescence*

Riss, T. L., Moravec, R. A., Niles, A. L., Duellman, S., Benink, H. A., Worzella, T. J., & Minor, L. (2016). Cell viability assays. In Assay Guidance Manual [Internet]. Eli Lilly & Company and the National Center for Advancing Translational Sciences.

Stoddart, M. J. (2011). Cell viability assays: introduction. In Mammalian cell viability (pp. 1-6). Humana Press.

Spectrum [Propidium iodide]