DAPI
DAPI or 4′,6-diamidino-2-phenylindole is a fluorescent stain that binds strongly to DNA. It is used extensively in fluorescence microscopy. Since DAPI will pass through an intact cell membrane, it may be used to stain both live and fixed cells.
For fluorescence microscopy, DAPI is excited with ultraviolet light. When bound to double-stranded DNA its absorption maximum is at 358 nm and its emission maximum is at 461 nm. (This emission is fairly broad, and appears blue/cyan.)[1] DAPI will also bind to RNA, though it is not as strongly fluorescent. Its emission shifts to around 500 nm when bound to RNA.[2]
DAPI’s blue emission is convenient for microscopists who wish to use multiple fluorescent stains in a single sample. There is fluorescence overlap between DAPI and green-fluorescent molecules like fluorescein and green fluorescent protein (GFP), or red-fluorescent stains like Texas Red, but using spectral unmixing or taking images sequentially can get around this.
Apart from labelling cell nuclei, the most popular application of DAPI is in detection of mycoplasma or virus DNA in cell cultures.
Because DAPI readily enters live cells and binds tightly to DNA, it is toxic and mutagenic. Care should be taken in its handling and disposal.
The Hoechst stains are similar to DAPI in that they are also blue-fluorescent DNA stains which are compatible with both live- and fixed-cell applications.
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