I've been able to activate mouse T cells to make CARs fine the last few months, but the T cells seem to not be activating as nicely and therefore are dying (even the non-transfected controls) and not expanding.
After two days of CD3/CD28+IL2 beads they barely expand and then after removal of beads and culturing for 2 days (with IL2) , they all die.
I have made new T cell media everytime - so I don't think that is the problem. I did start using new IL2 and new beads, but I have also tried multiple boxes and vials and still its poor.
Let me know if you have any suggestions on what to do or tips to improve as culturing mouse T cells has been a struggle for me the last year!
I have been experiencing the same problem with mouse CD4+ T cells. I have years of experience with activating/ expanding mouse T cells in culture. For the past few months, we are not able to keep them alive with a-CD3, a-CD28, IL-2. We have changed everything and the problem persists. Have you been able to solve your problem? Any faulty reagent we may be using in common?
I think we have found the culprits. They are most likely FBS and/or L-Glutamine. I recommend switching to Glutamax as it is more stable and testing different brands of FBS.