Culture with CSA reduces overall NK-cell fold expansion and results in higher percentages of CD56⁺CD16⁻ NK cells. (A) Fold expansion of NK cells cultured in IL-2 (100 U/mL) and IL-15 (10 ng/mL) in the presence of either vehicle control (EtOH, E) or increasing concentrations of CSA (0.1, 0.5, 1, and 5 μg/mL) after 7 days. Results are the average plus and minus the SD from 4 donors. Asterisk (*) indicates values that showed significantly less proliferation (P < .05) compared with vehicle controls using ANOVA with Tukey adjustment. (B) FACS plots from a representative donor showing staining for CD56 and CD16 before (left) and after 1 week of culture with IL-2 (100 U/mL) and IL-15 (10 ng/mL) and either vehicle control (EtOH) (middle) or CSA (1 μg/mL) (right). (C) The percentages of CD56⁺CD16⁻ and CD56⁺CD16⁺ NK-cell subpopulations after 7 days of culture in IL-2 (100 U/mL) and IL-15 (10 ng/mL) with or without CSA or vehicle (EtOH). Results of 14 individual donors are shown (gray dashed line) as well as averages (heavy solid line). Statistical analysis was performed using Wilcoxon singed rank test.