RE: Holding phagocytosis assays

We did fix the cells after terminating the assay. The cells were kept on 
ice after fixing till they were read on the flow. We used whole blood 
assays, so if you use the BD FACSlyse to lyseRBC, the cells get fixed too 
during the lysig step. You can work out conditions that suit you.



Indresh Kaur, Ph.D.
Coordinator, Flow Cytometry Lab
Department of Blood & Marrow Transplantation
713-563-4811 





"Waitumbi, John" <JWaitumbi@kisian.mimcom.net>

12/16/2004 01:38 AM




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Subject:
RE: Holding  phagocytosis assays



Why not stop the reactions by fixing the cells? Its as easy as keeping on
ice. 

John

-----Original Message-----
From: Julie Nelson [mailto:jnelson@uga.edu] 
Sent: Wednesday, December 15, 2004 5:10 PM
To: cyto-inbox
Subject: Holding phagocytosis assays

Hi ya'll,

One of my users poses the following question:

"In a phagocytosis assay, is there any point in the
procedure at which the reaction could be "put on ice"
(literally or metaphorically)?		 I ask this question
for a couple of reasons: the animals will be housed at
the Savannah River Ecology Lab (2.5 hours drive from
Athens), and I will be working with a large number of
animals, and even with help, I may not be able to
euthanize all the animals on the same day. 

I haven't found anything in the literature that
suggests that it would be okay to have any kind of
delays. The protocols include brief incubations, but
there seems to be no clear "holding point" in any of
the assays."

I imagine she'll be using either beads or FITC-labelled
bacteria.  Can you help?

Happy holidays and thanks in advance,

Julie
Julie Nelson
Research Coordinator
Flow Cytometry Facility
Center for Tropical and Emerging Global Diseases
University of Georgia