7-Aminoactinomycin D (7-AAD) is a fluorescent chemical compound with a strong affinity for DNA. It is used as a fluorescent marker for DNA in fluorescence microscopy and flow cytometry. It intercalates in double-stranded DNA, with a high affinity for GC-rich regions, making it useful for chromosome banding studies.

Applications

With an absorption maximum at 546 nm, 7-AAD is efficiently excited using a 543 nm helium–neon laser; it can also be excited with somewhat lower efficiency using a 488 nm or 514 nm argon laser lines. Its emission has a very large Stokes shift with a maximum in the deep red: 647 nm. 7-AAD is therefore compatible with most blue and green fluorophores – and even many red fluorophores – in multicolour applications. 7-AAD does not readily pass through intact cell membranes; if it is to be used as a stain for imaging DNA fluorescence, the cell membrane must be permeabilized or disrupted. This method can be used in combination with formaldehyde fixation of samples. 7-AAD is also used as a cell viability stain. Cells with compromised membranes will stain with 7-AAD, while live cells with intact cell membranes will remain dark. Viability of the cells in flow cytometry should be around 95% but not less than 90%.

Actinomycin D

The related compound actinomycin D is nonfluorescent, but binds DNA in the same way as 7-AAD. Its absorbance changes when bound to DNA, and it can be used as a stain in conventional transmission microscopy.

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