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trypan blue without hemocytometer?

Started by outremer, Mar 23 2010 02:04 AM

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2 replies to this topic

#1 outremer

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Posted 23 March 2010 - 02:04 AM

Hi, I´m a total novice in the cell lab. I wish to check cell viabity of osteoblasts cultured on glass surface. Is it possible to stain the cells with trypan blue and then observe the result in an ordinary light microscope and count the relative amount of dead cells?(not using hemocytometer which is the normal choice).

please share your thougts...

erik

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#2 Prep!

Am I me???!!!

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Posted 23 March 2010 - 03:22 AM

outremer, on Mar 23 2010, 03:34 PM, said:

Hi, I´m a total novice in the cell lab. I wish to check cell viabity of osteoblasts cultured on glass surface. Is it possible to stain the cells with trypan blue and then observe the result in an ordinary light microscope and count the relative amount of dead cells?(not using hemocytometer which is the normal choice).

please share your thougts...

erik

u can count it but u wont be able to calculate the viability percentage of the number of cells / some area etc as there are no accurate markings on a normal glass surface!!!
what ever u will get will be an approximation and it can well be far apart from the accurate reality!!!!
This is wat i blive... Its still open for suggestions!!

Support bacteria - They are the only culture some people have!!!
Cheers!!!

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#3 rhombus

Rhombus/Uncle Rhombus

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Posted 23 March 2010 - 03:38 AM

outremer, on Mar 23 2010, 10:04 AM, said:

Hi, I´m a total novice in the cell lab. I wish to check cell viabity of osteoblasts cultured on glass surface. Is it possible to stain the cells with trypan blue and then observe the result in an ordinary light microscope and count the relative amount of dead cells?(not using hemocytometer which is the normal choice).

please share your thougts...

erik

Very easy to do if you have a GRATICULE. This is an etched eye piece that fits into your microscope in the normal eye piece position. The graticule has a define area that can be used when counting cells that are adherent on glass and plastic. Just count more than 5 representative areas.....it is best to get another person to do this WHO DOES NOT KNOW THE EXPERIMENTAL PROTOCOL....this reduces the bias that can be introduced. The graticule area is known...you can adjust cell number for your particular flask/glass surface.

Hope this is useful

Kindest regards

Uncle Rhombus