How do you count Astain with DAPI?
Counterstaining protocol
- Dilute the DAPI stock solution to 30 nM in PBS.
- Incubate the specimen in the dark for 30 minutes at room temperature.
- Carefully remove the coverslip and rinse the specimen briefly with PBS or dH2O to remove unbound dye.
What is DAPI immunofluorescence?
DAPI is a classic nuclear counterstain for immunofluorescence microscopy, as well as an important component of high-content screening methods requiring cell-based quantitation of DNA content. DAPI is offered in powdered solid and aqueous solution forms.
Can I adding DAPI to a mounting media?
Yes, VECTASHIELD Antifade Mounting Medium with DAPI works well. I have used it before.
How do you Permeabilize a cell?
Permeabilizing the cells through methanol or acetone fixation, or with the use of a detergent, allows antibodies to pass through the cellular membrane and enter the cell. The most common reagent used for cell permeabilization is non-ionic detergent, Triton X-100.
How do you make DAPI dye?
DAPI stocks are usually prepared by simply dissolving the powder in sterile water, usually at 5 mg/mL. The solution may require gentle sonication to dissolve it all. A 5 mg/mL DAPI solution is stable for ~6 months at 2-6C, and much longer at -20C.
Is DAPI soluble in DMSO?
DAPI (hydrochloride) is soluble in organic solvents such as ethanol, DMSO, and dimethyl formamide (DMF), which should be purged with an inert gas. The solubility of DAPI (hydrochloride) in ethanol and DMF is approximately 0.2 mg/ml and approximately 3 mg/ml in DMSO.
How do I get glycerol mounting medium?
The following is the recipe for making glycerol mounting medium solution:
- di water ( bring it to the boiling point ): 60 ml.
- add gelatin slowly under constant stirring: 10 g.
- once fully dissolved, slowly add glycerin: 70 ml.
- add liquid phenol: 1 ml.
- aliquot preferably in 10 ml and store at room temperature until further use.