I would like to study Tcells which are CD4+CD25+FoxP3 separated from whole blood and compare them to those generated following stimulation. What is the best stimulant known to induce Treg cells in human PBMC?
Isolate the CD4 positive CD25 negative T cells and stimulate them with anti-CD3/CD28+ IL-2 +TGF-beta and incubate for upto 3-4 days
You can add retinoic acid in addition to cytokines mentioned by Asif to boost the generation of Tregs.
http://www.ncbi.nlm.nih.gov/pubmed/19717521
First you have to purify the ntregs from the PBMC. We use Milentyi isolation kits. Then you stimulate the cells in culture using CD3/28 beads (we use invetrogens t cell expansion beads at 3 bead:1 tcell ratio) or K562 with 64 and 86 cells (also called Kt64/86 cells) irradiated and loaded with anti-CD3 antibody at 1 Kt : 2 t cells. Then feed them IL-2 at 300IU/mL. To maintain the treg phenotype you will also need to treat them with Rapamycin.
http://www.ncbi.nlm.nih.gov/pubmed/21593401
Or if you want to induce t cells into tregs. You treat the isolated tcells with Rapamycin and TGF-B, with IL-2
http://www.ncbi.nlm.nih.gov/pubmed/21564534
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology