Do you use acetone as solvent for the stock solution? What concentration do you use (stock and in the actual staining) and how do you wash, if you do wash at all. I tried 1ug/ml in PBS (stock was in EtOH) for 30 min at RT but while washing, Nile red crashed out of solution and I lost most of my cells. Any suggestions on how to improve this?
Dissolve the 5ug Nile red dye in small quantity of ethanol/acetone (as samll possible) and make up the volume upto 1 ug/ml final concentration using the glycerol and mix it properly, use one drop of it for staining/ microscopy.
I would prefer dissolving the Nile Red in DMSO first and then make aliquots in the media as per you cell requirements.
Hope this helps..
Hm, so the actual concentration doesn't seem to matter that much. If you use glycerol, do you use this as a mounting reagent before microscopy? I would prefer a water based buffer to make my slides, to allow for live cell imaging in future experiments.
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