Embryonic Mouse Brain sort

Dear Fellow FLOWers:

I have a client who comes to me to sort disassociated embryonic mouse 
brains that have been transfected with GFP.  I am having real 
problems with the sorts as the samples contain large amounts of 
cellular debris as a result of the disassociation process. The client 
washes the samples multiple times trying to get rid of this debris 
with no success.

As a result, my sorter lines are being clogged, and I must stop the 
sort multiple times to backflush the sample lines(this last sort I 
had to stop 10 times).	I am certain the debris is causing the cells 
to clump and/or stick to the sample tubing.

I would be grateful to hear from anyone who may be doing similar 
samples with suggestions as to how we might stop this 
clumping/stickiness from disrupting the sort.

I am using a MoFlo with a 100 um tip running at 30 psi and am using a 
SmartSampler.  My stream stays remarkable stable during the sort, so 
I am thinking it is the sample tubing that is being clogged.  The 
sample is put thru a 35um mesh before sorting to eliminate large 
clumps.  The sample is kept at cold during the sort.

Thank you in advance for all the advice.

Sincerely,

Janet Dow
-- 
Janet Dow
Research Specialist and Manager
Flow Cytometry and Cell Sorting  Facility
North Carolina State College of Veterinary Medicine
4700 Hillsborough Street
Room C-309
Raleigh, NC 27606
(919)513-6443