If I want yo test the sensitivity of CD4+ T cells to a particular Ag. Is it correct to treat with antigen and stimulate with anti CD3/CD28 or do I have to use antigen presenting cell?
CD3/CD28 beads only mimic the stimulus provided by DC. But to test the response of CD4 T cells to a specific antigen, you should definitely pulse DC (or any other APC) with a specific antigen and coculture the T cells with these antigen pulsed DC. Read out systems can vary. You could do a Proliferation assay (FACS or thymidine incorporation) and analyse activation markers (FACS), test the cytokine production (ELISPOT or cytokine capture assays), etc. Hope it helps. All the best!