clumping and sieving

Ray Hicks rh208 at cam.ac.uk
Tue Sep 19 17:38:01 EST 2000


Hi Carolyn,

The death-related clumpy stickiness could be due to DNA from the dead cells
- try adding DNAse to prevent the re-clumping after filtration.

Ray

> From: Carolyn Jefferiss <prscmj at bath.ac.uk>
> Date: Tue, 19 Sep 2000 13:23:39 +0100
> To: Cytometry Mailing List <cytometry at flowcyt.cyto.purdue.edu>
> Subject: clumping and sieving
>
>
> Dear All,
>
> Has anyone tried using "Accumax" from TCS Cellworks at all in flow
> cytometry? It is supposed to be an anticlumping reagent, recommended
> for using for cell counting, which requires little or no Ca++ or
> Mg++. The cells are not suitable for culturing after disaggregation
> in this reagent. Anyone?
>
> My little bit on clumping on the sorting front;
> We sort cultured marrow cells which can be horribly clumpy and sticky
> (hence my wondering about Accumax) and we sort with a suspension at
> 10x10e6cells/ml having filtered the cells immediately before putting
> on the machine and the proportion of dead cells definitely affects
> the amount of clumpiness which develops. We don't put on more than
> 2ml at a time, and during a run take off the tube and give it a very
> gentle mix if it is really settling out, rather than using the buzzer
> on the machine. They can always get a second pass through a filter.
> The on-line thing sounds like a big faff if you get it clogged up so
> we've never done it. Since persuading everyone of the need to filter,
> before running each and every sample, we haven't had ANY blockages.
> (Famous last words).
>
> Carolyn Jefferiss
>
> Carolyn Jefferiss Ph. D.
> Pharmacy and Pharmacology
> University of Bath
> Claverton Down
> Bath BA2 7HY
>




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