dATP for TUNEL-FACS?

Ifor iforw at delphi.com
Tue Apr 18 11:04:46 EST 1995


I have recently begun using the TUNEL method to label the nicked ends of
DNA from apoptotic mouse thymocytes with biotin-21-dUTP using terminal
transferase.  After adding phycoerythrin-streptavidin as a second step
reagent, I am collecting data on the cells on a FACScan.  Some protocols
describing methods for TUNEL staining of cells for flow cytometry use a
labeled dUTP in combination with a molar excess of dATP (e.g. Sgonc et
al, Trends in Genetics, 10:41, 1994).
 
Can anyone tell me why adding dATP to the TdT reaction might improve
staining?  Does anyone have experience on whether this staining works
better with labeled dUTP alone vs. labeled dUTP and another unlabeled
dNTP?
 
Thanks in advance for any help.
Ifor Williams
E-Mail Address:  iforw at delphi.com
 



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