Propidium iodide (PI) is a popular red-fluorescent nuclear and chromosome counterstain. Since propidium iodide is not permeant to live cells, it is also commonly used to detect dead cells in a population.
PI binds to DNA by intercalating between the bases with little or no sequence preference. In aqueous solution, the dye has excitation/emission maxima of 493 / 636 nm. Once the dye is bound, its fluorescence is enhanced 20- to 30-fold, the fluorescence excitation maximum is shifted ~30–40 nm to the red and the fluorescence emission maximum is shifted ~15 nm to the blue, resulting in an excitation maximum at 535 nm and fluorescence emission maximum at 617 nm.
PI is widely used in fluorescence microscopy, confocal laser scanning microscopy, flow cytometry, and fluorometry.
High-performance nucleic acid stains
Nuclear counterstains provide highly selective nuclear staining with little or no cytoplasmic labeling and a choice of colors for multiplexing with other labels.
Some nuclear stains can also be used in cell cycle analyses, and others serve as dead cell stains, using high-content imaging and flow cytometry.
A fixed, permeabilized, and labeled muntjac skin fibroblast. Mitochondria were labeled with mouse anti–OxPhos Complex V inhibitor protein antibody and visualized using orange-fluorescent Alexa Fluor 555 goat anti–mouse IgGantibody. F-actin was labeled with green-fluorescent Alexa Fluor 488 phalloidin, and the nucleus was stained with TO-PRO-3 iodide (pseudocolored magenta).
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