Hello,
Quick question: in vitro, could I get a T cell activation/proliferation if I stimulate human PBMC with anti-CD3+IL-2?
thanks
vince
Yes you can stimulate Tc using anti-CD3 + IL-2 in the culture. For more potent stimulation of whole PBMC fractions, you could also consider adding anti-CD28. This can be done using aCD3/aCD28 coated plates or beads from commercial vendors as well.
-Benjamin
thanks for your reply. Will soluble aCD3 not induce cell death? ok, signals via TCR and CD28 would be optimal I guess.
Yes, the best option is CD3/CD28 beads use. A non-expensive alternative is PHA, It is a good T cells mitogen but induces high mortality.
You could also get memory cell specific T cell activation and proliferation through stimulation of cultures with a recall antigen. This is taken up by APCs, and presented to T cells in the context or MHC class II (or MHC class I), which causes them to proliferation. A commonly used antigen in PPD from M.tb I'm sure you could find others in the literature.
I use 0.5ug/mL anti-CD3 plate bound by coating and 2.0ug/mL anti-CD28 to the media. You can stimulate cells with PMA and ionomycin 20ng/mL and 1ug/mL, respectively. Stimulate with your specific antigen if you are assessing the proliferation against your specific antigen. Keep an unstimulated control for the comparison.
Please, correct me if I'm wrong!!
PHA on PBMC will activate T cells, but w/ or w/o apoptosis?
PHA on Jurkat will induce cell death or will it activate them (should I see CD3,28 internalisation?)
thanks
Hello,
Did anyone assess the seeding density of PBMC when activating them via aCD3/IL-2 "route"? thanks
I recommend you the easy to use Dynabeads™ Human T-Activator CD3/CD28 for activation of human T cells (Cat. no. 11131D).
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