I have been trying to activate my Jurkat T cells using CD3/CD28 DynaBeads and looking at activation markers such as CD69 and CD62L. However, there have been no signs of activation after multiple experiments.
Do you have any recommendations to improve my results, preferably without adding PMA/ionomycin?
Dear Richard Ii Dela Rosa
Please consider the following suggestions:
Improve your cell culture medium by switching to "complete" medium, i.e. RPMI with glutamine, pyruvate, non-essential amino acids, beta-mercaptoethanol and Hi-clone FBS as per
Article A role for the histone H2A deubiquitinase MYSM1 in maintenan...
may improve your cell culture and cellular activation results as per: For protocols please consider:
Article A Quantitative Assessment of Costimulation and Phosphatase A...
Article Quantitative Analysis of Phosphotyrosine Signaling Networks ...
Please also have a look at the following paper.
Article Mass-spectrometric identification and relative quantificatio...
As per Figure 5: CD62L is at best mildly upregulated, whereas CD69 does not seem to be upregulated at all following CD3/CD28 agonistic stimulation. They did culture for 24 hours in their experiment and used massspec as their readout, so therefore a time course might be a good experiment to do.
I hope that helps.
All the best & kind regards,
Michael
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