Looking for advice on pediatric samples

bunny bunny at cotleur.com
Tue Jun 21 16:14:13 EST 2005


Hello Flowers!

I'm looking for advice from someone/anyone who is currently analyzing 
small cell numbers by flow cytometry. (NOT low frequency, but very few 
events to begin with).

We are planning to start analyzing pediatric CSF samples in the near 
future. Since we are expecting very SMALL sample volumes with very FEW 
cells, I'm looking for advice on the best way to simultaneously 
quantify chemokine receptors, cytokine expression and cell phenotypes.

Is flow cytometry the best method in this case? With very few TOTAL 
cells, should we try a microscopy approach? Can someone point (or 
SHOVE) me towards the best method?

Our current equipment choices:
At the moment, our Flow core is only equipped with a Facscan and a 
LSR1. I could feasibly do 4 color  (with Viability on a 5th detector, 
using UV).
We are waiting to find out if we will be funded for an Aria. In any 
case, we could actually take samples across town to be run on an Aria. 
(I'm thinking in terms of maximizing # colors at this point).
We will also have a small amt of equipment money- enough for a 
FacsArray-type cytometer. Is there  an advantage (as far as running 
smaller samples) vs the traditional continuous stream? (We've removed 
the droplet containment on the LSR1 years ago, so we already fixed the 
problem with the DCM sucking the sample before we get the bar pulled 
over.  I regularly run 100ul samples without a problem).
Our institution also has a well equipped Confocal /Fluorescent 
Microscopy core.

Thanks for any input!

bunny




Bunny Cotleur, M.S.
Sr Research Technologist, Dept of Neurosciences
Scientific Consultant, Flow Cytometry Core
Cleveland Clinic Foundation
Lerner Research Institute, NC30
9500 Euclid Avenue
Cleveland, OH 44195
Lab: (216)444-1164
*********************************************
Caminante, no hay camino.      Se hace camino al andar.
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