Margitta-

I guess its not clear to me what you mean. The confocal image of FITC is
going to show you fluorescence. I don't see how you intend to do
densitometry on a fluorescent image. I, instead, you intend to measure the
intensity of the brightness of the images, it will be subject to the
issues of quantitative fluourescence that have been adressed here lately.
In answer to the Antifade, yes, absolutely use it. I would recommend the
Fluorgard product from BioRad since it does not appear to quench the FITC
signal. I would also look for fields under bright field, then collect the
images under fluorescence if at all possible or at least look for cells
under PI staining. If you look for bright cells under FITC and collect
them with FITC, you will be adding the error of photobleaching caused
while looking at the specimens and you will very much bias your data.

________________________________________________________________________________


Paul Goodwin
Image Analysis Lab
FHCRC, Seattle, WA

On Thu, 20 Feb 1997, Margitta Kampman wrote:

> I am planning to perform densitometry on tissue sections stained with
> FITC-conjugated antibodies. Should I use antifade mountant? Which one
> should I chose?
> Grateful for any suggestions,
> Margitta
>