Hello, i'm starting an assay soon about MSC (mesenchymal stem cell) and PBMCs coculture to see the inhibitory effect of normal MSC vs senescent MSC on PBMCs proliferation. I wonder if the use of anti cd3 / cd28 beads does not excessively activate proliferation preventing me from observing the desired inhibitory effect. is okt-3 enough to activate the proliferation, or there is another way to activate the PBMC properly for this assay ?
Thank you.
Hi Hicham:
1) The potency of CD3/CD28 beads from different suppliers is different. Read their manuals may give you some clue.
2) According to my understanding, only solid surface coated CD3 Ab can cross-link CD3 receptor, form cluster and immune synapse, and then trigger and activate T cells. Soluble IgM Ab may do, that's because their polymer structure. In your project, it is tricky to coat CD3 Ab and culture MSC adherent cells at same time. So CD3/CD28 beads look more feasible.
3) If you use CFSE for proliferation readout, sorted T cells may be better than PBMC. Since T cells just a subpopulation of PBMC, and it is difficult to synchronize them, the proliferation shift may be not obvious.
Thats' all i can think of, hope it helpful.
Regards
Shiqiu
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