I am not an immunologist, and I am trying to assay for IL-2, IFN-y, and TNF-a in vitro in primary cells, along with the surface markers CD69/25. Is treatment with anti-CD3/CD28 sufficient to induce cytokine production and surface marker expression due to T cell differentiation, or do I need to put the cells in certain/different conditions to induce these changes?
Hi Savannah West
agonistic stimulation with anti-CD3 (plate bound) and anti-CD28 is certainly sufficient to induce proliferation (which can be measured by CFSE dilution or IL2 in solution that can be assessed by ELISA). You can also measure cytokines like TNFa and IFNg by intracellular cytokine staining and activation markers like CD25, CD69 and CD44 by flow cytometry. For protocols and reagents for murine T cell activation and stimulation please check:
Article A role for the histone H2A deubiquitinase MYSM1 in maintenan...
Article Ubiquitin Specific Protease 21 Is Dispensable for Normal Dev...
All the best & good luck with your experiment,
Michael
anti-CD3/CD28 is sufficient for cytokine secretion and expression of receptors. You need to calibrate the doses and times which can affect things differently.
anti-CD28 is needed only if these are isolated T cellls
- Badges
- Science method
- Similar topics
- Biological Science
- Histology
- Staining