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CFDAse staining protocol
Chemicals:
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Carboxyfluorescein diacetate, succinimidyl ester (5(6)-CFDAse); dissolve
in DMSO to 5 mmol/L, aliquot in 50 µl and freeze in -20°C until use. |
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Bovine Serum Albumin (BSA) |
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PBS / 1% BSA |
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Phosphate Buffered Saline (PBS) |
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Wash
cells 3x in PBS to remove proteins.
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Aliquot cells at a
concentration of 1.107 cells/ml in a 15 ml tube.
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Dilute CFDAse stock 1 :
10,000 to 0.5 µmol/L in PBS from the 5 mmol/L stock.
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Spin cells down, remove supernatant, and add 1
ml 0.5 µmol/L CFDAse solution per 1.107 cells. Resuspend cells well.
Incubate 5 minutes at RT.
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Wash the cells 2x with PBS / 1% BSA at 4°C to
stop the labelling.
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Cells can be used for assay now.
NB. Due to high intensity of the signal
after labelling. This can be checked best the next day.
NB2. The population of undivided cells should be identified by culturing
a portion of the same cells for the same period without activation.
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Protocol CFDAse
