Anti cd3ε fitc clone 145.2c11

Cell-free co-culture supernatants were harvested at the indicated times. IL-2 and IFNγ were determined by immunoassay. Antibody pairs were purchased from Biolegend. Cytokine levels were determined using streptavidin-europium and enhancement solution (both from Perkin Elmer) and detected on a Victor 1420 multilabel counter (Perkin Elmer). Day 1 or 6 BMDC and OT-II T-cell co-cultures were harvested and stained with anti-CD3ε-FITC (clone 145.2C11; Biolegend), anti-CD4-PerCP (clone RM4-5; Biolegend), anti-CD69 (clone H1.2F3; Biolegend), anti-CD25 (PC61; Biolegend) and Fixable Viability Dye eFluor 506. Cells were fixed in 1% PFA in PBS. Proliferating and activated T-cell populations were determined gating on live, singlet, CD3⁺, CD4⁺ cells and by CTV dilution.

Cell-free co-culture supernatants were harvested at the indicated times. IL-2 and IFNγ were determined by immunoassay. Antibody pairs were purchased from Biolegend. Cytokine levels were determined using streptavidin-europium and enhancement solution (both from Perkin Elmer) and detected on a Victor 1420 multilabel counter (Perkin Elmer). Day 1 or 6 BMDC and OT-II T-cell co-cultures were harvested and stained with anti-CD3ε-FITC (clone 145.2C11; Biolegend), anti-CD4-PerCP (clone RM4-5; Biolegend), anti-CD69 (clone H1.2F3; Biolegend), anti-CD25 (PC61; Biolegend) and Fixable Viability Dye eFluor 506(eBiosciences). Cells were fixed in 1% PFA in PBS. Proliferating and activated T-cell populations were determined by gating on live, singlet, CD3⁺, CD4⁺ cells and by CTV dilution.