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Javier, 
There has been much discussion about this recently, and I am not sure that there was any positive response (correct me if I'm wrong), concerning blue-emitting dyes. I have tried them again myself recently using conventional microscopy, with no luck, even at 10x-20x [ ] of what I would normally use other Alexa Fluors (AF350, Pacific Blue, Marina Blue, etc),  along with the ProLong Gold antifade that is supposed to retard quenching in these samples. I have talked to a few others who have also experienced the same poor results. 

If anyone has had good luck with these, or figured out the trick to achieving good signal, I would also love to find out as well.

Good luck, 
Melissa


Melissa A. Gonzalez
R&D Cell Genesys
500 Forbes Blvd. 
South San Francisco, CA 94080 

-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]]On
Behalf Of F Javier Diez Guerra
Sent: Monday, May 08, 2006 7:25 AM
To: [log in to unmask]
Subject: best blue-emitting dye?


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Dear colleagues,

I need to visualize four fluorophores in fixed samples using a conventional 
fluorescence microscope that has filters to detect FITC/GFP/Alexa-488, 
Alexa-594/TxRed, Alexa-647/Cy5 and DAPI/Hoechst.

I would like to hear opinions and experiences with blue-emitting 
fluorophores such as alexa-350, alexa-405, coumarin, etc in terms of 
brightness (extinction + QY), photostability, etc that could be used with 
the DAPI filter set.

Thanks.



F Javier Diez-Guerra, PhD
Profesor Titular
Centro de Biologia Molecular Severo Ochoa
Facultad de Ciencias, Universidad Autónoma
Ctra Colmenar Viejo Km 15
Cantoblanco, 28049 Madrid
SPAIN

phone:  +34 91 4978051
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