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Nile Red and Cell Harvesting

Nile Red and Cell Harvesting - Flow Cytometry Forum

Nile Red and Cell Harvesting - Flow Cytometry Forum. Discuss FACS cell sorting, flow cytometry, analysis and cell staining.


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Old 12-03-2010, 05:59 PM
Pipette Filler
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Default Nile Red and Cell Harvesting



Hi im working on my thesis about hepatic steatosis...

I usually (SOP) overload my cells with palmitic and oleic acid...but just recently when i ran it through flow cytometry it turns out that my cells apparently were 'not stained'. I use nile red at 100ng/mL (10mg/10mL DMSO, taken 5uL dilute to 50mL = 100ng/mL); i incubate my cells at 37 degrees for 10 minutes in the nile red solution; around 2mL of 100ng/mL in 3 million cells/mL. There is absolutely no sign of stained cells in the flow cytometry experiment...please help?


in this experiment my superior told me that neutralizing trypsinized cells with 10%FBS in PBS is okay. is this the problem?


theoretically speaking if i prepared the stain as deascribed there should be some indication in the flow cy experiment but there isnt...can anyone help me with this?


URGENT....


thanks
evilhamster
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