Nov 24, 2021
DAPI reconstitution and viability staining
This protocol is a draft, published without a DOI.
- 1University of Newcastle-upon-Tyne
Protocol Citation: Emily Stephenson 2021. DAPI reconstitution and viability staining. protocols.io https://protocols.io/view/dapi-reconstitution-and-viability-staining-b2b3qaqn
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: November 24, 2021
Last Modified: November 24, 2021
Protocol Integer ID: 55387
Abstract
For the reconstitution of DAPI to make a stock solution and dilution for a working solution.
Details of viability staining volumes also provided.
Materials
DAPI (Sigma Aldrich, D9542-10mg)
Filter-sterilised dH2O
Sterile 1x PBS
Reconstitution
Reconstitution
Add 2ml of water to the 10mg vial of DAPI to make a 14.3mM (5mg/mL) stock solution. Vortexing may be required if the powder does not dissolve with pipetting.
(Note: DAPI will not dissolve in PBS)
Add a label with the date of reconstitution and store this stock solution at 4 degrees C for 6 months.
Working Stock
Working Stock
Add 420uL DAPI stock to 20mL PBS to make a working solution of 300uM.
Store in the dark at 4 degrees C.
For staining
For staining
For immunofluorescence, add to fixed cells at a concentration of 300nM (i.e. 1:1000 dilution for 5 mins then wash cells in PBS 3 times.
For flow cytometry, add to live cells at a concentration of 3mM (i.e. 1:100)