T cell Purification

[rjanssen_imm at opal.tufts.edu]

In article <36kj8j$qu at netnews.upenn.edu>, dfonseca at mail1.sas.upenn.edu
 (Dina Fonseca) writes:
> : The only T cell that would be unresponsive to PHA would be an anergized
> : one. Is this what you're asking for?
> 
> : Do you want helper CD4s or cytotoxic CD8s or both?
> : You could select for these markers. As far as negative selection,
> : you're in for a long experiment.  There's over 200 other non-T cells
> : you'll have to negatively select out for removal.
> :  How about + selection for CD40-ligand?
> : --                          ^^^^^^^^^^^
> :                                              psycler at netcom.com
> I assume these are human T cells.
> I did'nt know resting human T cells expressed CD40L.
> Is this true?
> A Bhandoola
>

Resting T cells do, to my knowledge, not express CD40L. And when activated,
onlly the T helpper population will express the ligand. 

Two hundred non-T cell populations is probably a way of speaking ;-). You could
 try magnetic beads coated with CD14, CD19, CD16 and CD56 binding monocytes, B 
cells and NK cells  from a mononuclear cell preparation. You will lose however the
CD56 positive T cells ! Another possibility is the use of a flow cytometer
using the antibodies above. This is however very costly, but gives good
results. Using the ELITE from Coulter (NO CONNECTIONS!) I was able to get about
99% pure T cells. Th best purification was positive selction with magntic
beeads and OKT3 antibody. I did not check for activation, but it probably does
activate your cells, despite the rapidness of the method.

Good luck
Richard Janssen