DAPI is a popular nuclear counterstain for use in multicolor fluorescent techniques. Its blue fluorescence stands out in vivid contrast to green, yellow, or red fluorescent probes of other structures. When used according to our protocols, DAPI stains nuclei specifically, with little or no cytoplasmic labeling. The counterstaining protocol is compatible with a wide range of cytological labeling techniques—direct or indirect antibody based detection methods, mRNA in situ hybridization, or staining with fluorescent reagents specific for cellular structures. DAPI can also serve to fluorescently label cells for analysis in multicolor flow cytometry experiments.
The excitation maximum for DAPI bound to dsDNA is 358 nm, and the emission maximum is 461 nm. DAPI can be excited with a xenon or mercury-arc lamp or with a UV laser. DAPI may be used in flow cytometry systems utilizing UV excitation sources.
To make a 5 mg/mL DAPI stock solution (14.3 mM), dissolve the contents of one vial (10 mg) in 2 mL of deionized water or dimethylformamide (DMF).
Note: DAPI is not very soluble in phosphate-buffered saline (PBS).
For long-term storage, the stock solution can be aliquoted and stored at -< –20°C. For short- term storage, the solution can be kept at 2–6°C, protected from light. When handled properly, DAPI solutions are stable for at least six months.