I have activated human T cells in vitro (with CD3/CD28 for 1-2 days) and then stimulated them shortly with rhTNF (0-30 min). After CD3/CD28 activation, basal p-STAT3 levels incresead compared to the control T cells. However, upon TNF stimulation, preactivated T cells showed decreased p-STAT3 MFI. On the other hand, unactivated T cells had only slightly increased p-STAT3 levels. Any comments?
Are your FACS data really convincing enough so that you can make those asumptions?
I would privilege Western Blot instead of intracellular FACS in order to make sure the signal that I look at really is coming from p-STAT3.
Also please consider the fact that activated T cells will endogenously produce TNF and that they produce IL-2 (which induces some STAT3 signaling), while unactivated T cells kept in culture for 2 days start/are dying (which could lead to a poor response to TNF).
- Badges
- Science topic
- Similar topics
- Comment