Anti mouse cd4 fitc mab

T cells were isolated from the spleen and inguinal lymph nodes of the iTreg-treated, iTreg_mtDC-treated, and CIA mice on day 49 after the primary immunization. These cells were incubated with an anti-mouse CD4-FITC mAb, fixed, permeabilized, stained with anti-mouse FoxP3-PE, IL-17-PE, or isotype control mAbs (BD Biosciences Pharmingen), and then assessed by flow cytometry.

As previously described [12 (link)], CD4⁺CD25⁻T cells were purified from the spleens of D1 mice using a CD4⁺CD25⁺ regulatory T Cell Isolation Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) and were subsequently stimulated with anti-CD3/CD28 monoclonal antibodies (mAbs, 50 ng/mL, PeproTech) in the presence of TGF-β1 (5 ng/mL, PeproTech) for 5 days. The T cells were incubated with anti-mouse CD4-FITC mAb, fixed, permeabilized, and stained with anti-mouse FoxP3-PE or isotype control mAbs (BD Biosciences Pharmingen). And the suppressive activity of these cells against T cell proliferation was examined using an in vitro suppressive assay. Approximately 3 × 10⁶ cells were transferred to CIA mice at 25 days after the first CII immunization.