PE-Cy7 and compensation problems

Fischer, Randy (NIH/NIAMS) [E] fischer1 at
Sat Apr 14 21:41:48 EDT 2007

Hi Virginia,

This sounds like multiple problems.  Starting with the second problem, the PE and PE-Cy7,
this should not be an issue unless your tandem is bad.	We, and others, routinely use PE
and PE-Cy7 without any issues.	This really sounds like your PE-Cy7 has become
dissociated with the PE now being the only fluorochrome left.

With the problem of APC in your PE-Cy7, how much spillover?  I would expect very little
to none normally for two reasons-different light paths via the fiber optics from the two
different lasers and very different emission curves.  Off the top of my head this sounds
like something misaligned-have BD check it out.

By the way, I have used the BD 34 PE-Cy7 and not had problems.

Randy Fischer

-----Original Message-----
From: Virginia Portillo [mailto:Virginia.Portillo at]
Sent: Thu 4/12/2007 11:48 AM
To: cyto-inbox
Subject: PE-Cy7 and compensation problems

I would like to do some multicolor experiments in a FACS Aria using the folowing

Rh123 (FL-1)
Hst 33342
CD34 PE-Cy7 tandem dye
monoclonal Ab detected by a APC- conj. sec Ab

We have tryed a similar experimet in the past and had some problems using the
CD34 PE-Cy7 Ab such as:
-APC leaking on the PE-Cy7 channel
-when using PE together with PE-Cy7, could not compensate either.

All comercial Abs I use are BD so they do not differ on fluorochrome:protein
molar ratios.

I am thinking on changing my CD34 or secondary Abs but for example the emission
of CD34 Cy5.5-PerCP would overlap with APC.

I would appreciate your comments on using tandem dyes in particular PE-Cy7
conjugates and/or any suggestions about using different color combinations on
my experiment to avoid these problems.

Many thanks!



Dr Virginia Portillo
The University of Manchester
Manchester Interdisciplinary Biocentre
131 Princess Street
M1 7DN

Tel: 0161 306 5820
Fax: 0161 236 0409

More information about the Cytometry mailing list