We routinely do CFP/YFP on our Facstar Plus. I use the 457 line on the 2nd laser to excite the CFP; this works just fine. I got a 470/10 nm bandpass filter from Omega which I put in FL3(2) [== P5] with the 505SP dichroic. This improved matters considerably as the standard 485 bandpass filter allows too much interference from the first laser when the drop drive is turned on. That said.... we tried CFP and GFP together and it _didn't_ work well. The problem is that (E)GFP excites quite nicely at 457 and also emits an appreciable amount at 470, so the CFP signal was spoiled by the GFP. I would try to get your users to use YFP instead of GFP, since the CFP/YFP color combo works very nicely. You can get all the spectra for Clontech's fluorescent proteins from their web site: Try: http://www.clontech.com/archive/OCT99UPD All the best -- David On Sun, 07 May 2000, you wrote: > I have a user who wishes to run Clontech EGFP and ECFP together; we > can run EGFP O/K but we have not yet tried ECFP & I'm dubious.... > > Has anyone used ECFP (cyan) in a flow cytometer? > If so, what laser & wavelength excitation? > > The Clontech website says excitation is 433 with a secondary > absorption peak at 453. > No spectra shown, so I don't know how broad the absorption curve > might be. > We don't have any 433 line available, would the argon 457 line work? > > Any ideas &/or experience? > > Thanks, Joseph. > -- > Joseph Webster, Flow Cytometry Facility > Centenary Institute, Sydney AUSTRALIA.
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