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There is another possibility - use lifetime imaging to distinguish
between dyes with similar spectra but different lifetimes.  Carlsson, K.
and Liljeborg, A., 2009.  Confocal fluorescence microscopy using
intensity-modulated multiple-wavelength scanning (IMS): evaluation of
results from spectral and lifetime imaging. Proceedings of SPIE 3261, 30

                                     Guy

Optical Imaging Techniques in Cell Biology
by Guy Cox    CRC Press / Taylor & Francis
     http://www.guycox.com/optical.htm
______________________________________________
Associate Professor Guy Cox, MA, DPhil(Oxon) 
Australian Centre for Microscopy & Microanalysis, 
Madsen Building F09, University of Sydney, NSW 2006 

Phone +61 2 9351 3176     Fax +61 2 9351 7682
             Mobile 0413 281 861
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-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]]
On Behalf Of Olivier Bardot
Sent: Thursday, 10 November 2011 8:52 PM
To: [log in to unmask]
Subject: Re: 5th colour?

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Hello
You can use Chromeo 494. It has a long stokes shift (Em 628nm) that
allow you 
to excite two dyes (any 488 et chromeo494) with the same 488 excitation
laser.
Hope this help.

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