Figure 1. Frequency and Yield of CD7⁺CD5⁺ Pro-T Cells After 14 Days of Culture

CB-derived CD34⁺ cells (freshly isolated or frozen) were cultured for 14 days in StemSpan™ SFEM II containing Lymphoid Progenitor Expansion Supplement (Catalog #09915) on plates coated with Lymphoid Differentiation Coating Material (Catalog #09925). Cells were harvested and analyzed for CD7 and CD5 expression by (A) flow cytometry. The (B) average frequency of viable CD7⁺CD5⁺ pro-T cells on day 14 was 70%, with ~200 CD7⁺CD5⁺ cells produced per input CD34⁺ cell. Shown are means with 95% confidence intervals (n = 33).

Figure 2. Frequency and Yield of CD4 ISP and CD4⁺CD8⁺ DP Cells After 42 Days of Culture

CB-derived CD34+ cells (freshly isolated or frozen) were cultured with the StemSpan™ T Cell Generation Kit (Catalog #09940) for 42 days and (A) analysed by flow cytometry for the expression of CD4, CD8, CD3 and TCRαβ. The (B) frequency and (C) yield of CD4 ISP, double-positive (CD4⁺CD8⁺) and CD3⁺TCRαβ⁺-expressing double-positive cells (CD4⁺CD8⁺CD3⁺TCRαβ⁺) are shown. On average, 38% of the total viable population were DP (CD4⁺CD8⁺), of which 35% co-expressed CD3 and TCRαβ. The yields of total DP cells and CD3⁺TCRαβ⁺ DP cells per input CD34⁺ cell were ~23,000 and ~9,000, respectively. Shown are means with 95% confidence intervals (n = 31).

Figure 3. Frequency and Yield of CD8 SP T Cells After 49 Days of Culture

DP cells were further matured into CD8 SP T cells by culturing for an additional 7 days in StemSpan™ SFEM II with T Cell Progenitor Maturation Supplement (Catalog #09930), IL-15 (Catalog #78031) and ImmunoCult™ CD3/CD28/CD2 T Cell Activator (Catalog #10970) on coated plates. On day 49, cells were (A) analyzed by flow cytometry for the expression of CD3, TCRαβ, CD4 and CD8. The (B) frequency and yield of CD3⁺TCRαβ⁺-expressing cells and their subsets are shown. On average, 54% of the CD3⁺TCRαβ⁺ cells were DP (CD4⁺CD8⁺) and 38% were CD8 SP (CD4^-CD8⁺). The average yield of CD8 SP T cells per input CD34⁺ cell was ~6,000. CD3⁺TCRαβ⁺ CD4 SP (CD4⁺CD8^-) T cells were detected at very low frequencies (data not shown). Shown are means with 95% confidence intervals (n = 12).