Caspase-3

From:Colleen Forster

Hi Collene:

    Thanks for the information. Did you send it to the Histonet and/or 
the original poster?

Geoff

Colleen Forster wrote:

Hi Geoff,

I stain para fixed mouse tissue with Caspase-3 with good results. The 
real key to this stain is to have a CONTROL KNOWN TO HAVE APOPTOTIC 
CELLS. I have a transgeneic mouse control that I use and the control 
stains beautifully. However, even then there are FEW positive cells. 
Another suggestion was to use the intestine as the cells in the 
fingerlike crypts do change over at a rapid rate. I have not done my own 
trials yet. I have collected the tissue.

I am using the Promega polyclonal Caspase-3 at 1:800. I run the slides 
to water, steam with a citrate buffer for 30 min followed by a 10 min 
cool down, rinse and stain. I leave my primary on overnight at 4 
degrees. I follow with detection from SignetLaboratories, a biotinylated 
secondary, streptavidin third, develop with DAB. As a rule you will not 
see large numbers of positively stained cells. The staining pattern is 
nuclear.

Colleen Forster
U of MN




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