Ghost Dye™ viability dye (see http://www.tonbobio.com/ghostdyes for available formats)
Other Materials Required
- 1X PBS (azidefree, protein/serumfree)
- Flow Cytometry Staining Buffer (Stain Buffer) (1X PBS with 2% FBS, 0.09% Na-Azide)
- Remove Ghost Dye vial from freezer and allow to equilibrate to room temperature.
- Quick spin Ghost Dye vial before opening.
- Prepare azide and protein/serum-free 1X PBS for labeling procedure.
- Wash cells twice in 1-2 mL azide and protein/serum-free 1X PBS. Spin at 300-400 x g for 5 minutes at room temperature and decant supernatant.
- Resuspend cells to a concentration of 1-10 x 10^6 /mL in azide and protein/serum-free 1X PBS.
- Add 1 μL of Ghost Dye solution for each 1 mL of cell suspension and vortex immediately. Note: Ghost Dyes are formulated in DMSO, pipet carefully and slowly.
- Incubate for 30 minutes at 2-8°C protected from light.
- Wash cells 1-2 times with 1-2 mL Stain Buffer. Washing with a protein containing buffer allows removal of any unreacted dye prior to staining with fluorescent antibodies.
- Cells can be subsequently stained, fixed and permeabilized according to user protocol.
Note: Cells labeled with Ghost Dyes can be cryopreserved for later use or used in intracellular staining protocols without any loss of fluorescence intensity.