Figure 1. ImmunoCult™ Mouse Treg Differentiation Supplement Produces CD4+CD25+FOXP3+ Cells Under Treg Polarizing Conditions

Naïve CD4+ T cells were isolated from mouse splenocytes using the EasySep™ Mouse Naive CD4+ T Cell Isolation Kit (Catalog #19765), activated with plate-bound anti-CD3 and soluble anti-CD28 and cultured in medium alone (Non-polarizing cultures), or in medium supplemented with ImmunoCult™ Mouse Treg Differentiation Supplement (Polarizing cultures) for 6 days. Cells were subsequently stained with anti-CD4, anti-CD25, anti-FOXP3 and a viability dye and analyzed by flow cytometry. Shown is the expression of CD25 and FOXP3 back-gated on viable, CD4+ cells from non-polarized (A) or polarized cultures (B). The mean proportion of CD4+FOXP3+ cells is significantly higher in cells cultured in ImmunoCult™ Mouse Treg Differentiation Supplement (91 ± 2%) compared to non-polarized cells (2 ± 0.4%) (p < 0.001; n = 14). Data from experimental groups were compared using a paired T-test.

Figure 2. ImmunoCult™ Mouse Treg Differentiation Supplement Expands Viable CD4+ Cells Under Treg Polarizing Conditions

Naïve CD4+ T cells were isolated from mouse splenocytes using the EasySep™ Mouse Naive CD4+ T Cell Isolation Kit (Catalog #19765), activated with plate-bound anti-CD3 and soluble anti-CD28 and cultured in medium alone, or medium containing ImmunoCult™ Mouse Treg Differentiation Supplement for 6 days. A 3.6-fold expansion of total nuclear cells (TNC) ensues in cultures polarized with ImmunoCult Mouse Treg Differentiation Supplement (purple column), which is significantly higher compared to non-polarized cultures (grey column). (Data represent the mean ± SEM, n = 14, ***p < 0.001. Data from experimental groups were compared using a paired T-test).