DAPI Fluorescence Counterstain Protocol




DAPI Counterstain Solution:

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. DAPI stains nuclei specifically, with little or no cytoplasmic labeling. The following protocol is used for staining of tissue sections or for staining cultured cells on slides.

DAPI Stock Solution (5mg/ml or 14.3 mM):


DAPI (Molecular Probes, Cat# D-1306) ---------------- 10 mg

Dimethylformamide (DMF) ------------------------------ 2 ml

Mix to dissolve and it may take sometime to completely dissolve.

Aliquot and store in –20 ºC.


DAPI Working Solution (100ng/ml or 300 nM in PBS):


DAPI stock solution -------------------------------------- 2 ul

PBS -------------------------------------------------------- 100 ml

Store this solution at 4 ºC in brown bottle or wrapped with aluminum foil to protect from light (to achieve stronger staining, use 4 ul stock solution or reduce PBS amount to 50 ml).


Incubate sections in dark for 30 minutes at room temperature.


Staining Pattern


Nuclei will be stained bright blue.


Suggested Use


Counterstain for immunofluorescence when green (FITC) or red (Texas Red) fluorescent marker is used.