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Figure 1. Typical EasySep? Mouse Na?ve CD4+ T Cell Isolation Profile

Starting with mouse splenocytes from an uninfected mouse, the na?ve CD4+ T cell content (CD4+CD44lowCD62Lhigh) of the isolated fraction typically ranges from 87.6 - 95.6%. In the above example, the purities of the start and final isolated fractions are 14.3% and 94.7%, respectively.

Figure 2. CD4+ Mouse Splenocytes Labeled with PE-Conjugated, Anti-Mouse CD62L (L-Selectin) Antibody, Clone MEL-14

(A) Flow cytometry analysis of C57BL/6 mouse splenocytes labeled with Anti-Mouse CD62L (L-Selectin) Antibody, Clone MEL-14, PE (Catalog #60109PE; filled histogram) or Rat IgG2a, kappa Isotype Control Antibody, Clone RTK2758, PE (Catalog #60076PE) (solid line histogram). (B) Flow cytometry analysis of mouse splenocytes (gated on CD4+ cells) processed with EasySep? Mouse Na?ve CD4+ T Cell Isolation Kit (Catalog #19765) and labeled with Anti-Mouse CD62L (L-Selectin) Antibody, Clone MEL-14, PE and an anti-mouse CD44 antibody, FITC. (C) Labeling of the Start splenocytes prior to cell isolation.

Figure 3. ImmunoCult? Mouse Treg Differentiation Supplement Produces CD4+CD25+FOXP3+ Cells Under Treg Polarizing Conditions

Na?ve 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 (Catalog #10957; 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 ¡À 2%) compared to non-polarized cells (2 ¡À 0.4%) (p < 0.001; n = 14). Data from experimental groups were compared using a paired T-test.

Figure 4. ImmunoCult? Mouse Treg Differentiation Supplement Expands Viable CD4+ Cells Under Treg Polarizing Conditions

Na?ve 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 (Catalog #10957) for 6 days. A 3.6-fold expansion of total nuclear cells (TNC) occurs 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 ¡À SEM, n = 14, ***p < 0.001. Data from experimental groups were compared using a paired T-test).