BrDU proliferation assay

Olav Engebraaten test
Thu Feb 23 12:00:07 EST 1995


In article <1995Feb23.173459.7706 at ucsvc.ucs.unimelb.edu.au>,
chris at austin.unimelb.edu.au wrote:

> Has anyone had experience with the BrDU labelling assay
> for cell proliferation?  I have attempted to use it in 
> a manner analagous to the continuous labelling of cells
> with tritiated thymidine.  This a simple method giving
> much information of cell cycle time etc.  However, 
> rather than getting a simple increase in labelling over
> time with an eventual plateau when all cells are 
> labelled, I obtained a sine curve with two peaks, the 
> second less than the first.  I would be glad of any 
> explanations (or even half convincing fictions!).
> 
> Thanks in advance,
> Chris Bradley


I have worked somewhat with BrdU labeling visualized both by flow cytometry
and immunocytochemistry, but have never encountered the problem you
describe. However, I have almost always used a BrdU concentration of 10 uM
and used a concomitant addition of 10 uM of deoxycytidine in the medium. I
have primarily been interrested in obtaining the number of replicating
cells, and only done pulse labeling of cells for 1 hr. 
I know that you may run into toxicity problems using BrdU. In case of
trouble, you should try to run some assay determining the safe level of
BrdU to use on your cells (clonogenicity assays or even growth curves).
I wish you good luck!

Best regards

Olav Engebraaten

Dept.of tumor biology
The Norwegian Radium Hospital
Oslo, Norway



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