Filters and Cy5

Dave Coder dave at nucleus.immunol.washington.edu
Fri Feb 4 11:19:10 EST 1994



Begin forwarded message:

Date: 03 Feb 94 16:38:51 EST
From: Marc.Langweiler at Dartmouth.EDU (Marc Langweiler)
Subject: Re: Sci Fi (ve) mystery
To: dcoder at u.washington.edu
X-Status: 

======
--- You wrote:
Glad it helped. I'd look at the transmission properties of the 660LPDC.  

Depending how sharp the rise is around 660nm, you may reflecting some Cy5  

emission since the emission peak of Cy5 isn't much beyond 660  (ML - 667 ??).
--- end of quoted material ---

And them Marc wrote:

Dave,
  Thanks for your repost.  In fact, I have gotten Coulter to send me an
entirely new filter set.  You were absolutely right about the 660LPDC - it
looked really different than another of the identical filter (though we
didn't see any difference upon substitution).  I also contacted a "local"
filter vendor (Omega Optical in Brattleboro, VT) who suggested we might be
better served using a 670 BP in lieu of our 645 LPA, an opinion echoed by
other respondents.
.....
============
Marc,

When we first started using Cy5 somewhere in 1988 or 1989 (when Alan Waggoner  
had just gotten it out of the lab) I had Omega make a 670DF35 filter which  
worked very well. To get as much of the Cy5 signal as possible, we picked the  
red emission off first by using a 650SP DC (also from Omega). We could excite  
Cy5 (direct conjugates or Cy5-StrAv) with a 35mW HeNe laser (632nm) on an EPIC  
753 with jet-in-air flow cell and get very good results. (FITC or PE  
sensitivity can suffer since they are now pickup farther down the optical path  
several other filters later. You play toward your dimmest probes and try to  
maximize their detection.)  


Omega should still offer the filters. Another source is Chroma Technology  
formed by people who left Omega:

Chroma Technology
RR 6 Box 9E
Brattleboro, VT  05301
(802) 257-1800
(802) 257-9400 fax

Paul Millman is with Chroma and he's the one who got me the filters originally.

Dave




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