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what is the measurement of Oil red O staining


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#1 meredith

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Posted 06 April 2010 - 06:26 AM

Hi,
I'm doing oil red O staining of macrophages,but what is the measurement of ORO staining?
The number of cells stained by ORO?
The fluorescense intensity of ORO?
Or comparing the red area in cells?

Thanks!

#2 bob1

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Posted 06 April 2010 - 04:23 PM

It all depends on what you are looking at... sections of tissue? plated cells?

#3 jah

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Posted 06 April 2010 - 04:45 PM

Oil red O stains lipid droplets. What you measure, As Bob1 said, depends on tissues vs. cells. Assuming cells, you are probably best off taking a direct measure of ORO bound. To do this you need to make sure you wash all of the un-bound stain (background) from the cells. You can then elute the ORO in isoproanol, and quantify in a spectrophotometer; peak absorbance is 518nm, though we use a 492nm filter. For comparison between replicate experiments, you can formulate a standard curve of ORO diluted in Isopropyl alcohol.

#4 meredith

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Posted 07 April 2010 - 01:58 AM

Thank you, bob1 and Jah.

I am going to compare the degree of foam cell formation between 2 groups of mouse peritoneal macrophages.

As I know,there are 2 measurements for this:
1) ORO staining
2) Quantitation of intracellular contents of cholesterol

Are there any else ways?

For oil red O staining,is OD/mg protein suitable?
or fluorescence intensity?




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