I would like to study the influence of different dendritic cells on the polarization of T cells in vitro. However I have a very poor survival of DCs (especially pDCs) when I culture them in vitro after MACS separation. I already tried rIL-3 for the pDCs but it didn't really help...
Hey,
I think one of the problems is, that the MACS protocol is not very short and also the steps are stressful for the cells.
Maybe you could already optimize the MACS protocol to stress the cells less.
One important point could be that you negative MACS.
We are just differentiating DCs from the bone marrow but maybe also in your case GM-CSF would help.
Maybe that helps.
all the best,
Sebastian
Thanks for the answer but unfortunately I really need primary DCs isolated from my tissues of interest in order to look at their induction of Treg ... GM-CSF would alter the phenotype of DCs (it's pro-inflammatory :-( and I want to test DCs from an anti-inflammatory / tolerogenic environment )
Hi, Do you culture the pDCs/DCs for some time such as 24h before coculture with naive T, or conduct the coculture immediately after the purification? Besides, check your IL-3.
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