Proliferation Assay Protocol to Measure T Cell Responses

Similar protocol was used by Rosenthal and Shevah to define MHC II restriction.

1. Protein in adjuvant in injected into a mouse (e.g. H-2b). This immunizes the mouse.

2. Allow 2-3 weeks for T cell production response.

3. Isolate T cells (depletion of APC)

4. Add irradiated spleen cells plus antigen (this can be either a protein or peptide)

5. Incubate for 48 hours

6. Add 3H-thymidine

7. Incubate for 6 hours

8. Measure 3H uptake into DNA (lyse cells in water, collect DNA through a filter)