In vitro differentiation of cord blood CD4⁺ cells for Th17 gene expression profiling. (A) IL17A secretion after 72 hours of culture. The detection was done directly from the culture supernatant with fluorescent beads. The values are normalized with the number of living cells determined based on cell size and granularity detected with flow cytometer. The data shown are the average of 6 cultures with the SEM and statistical significance was determined with the Student t test. (B) The expression of IL17F and RORC in the cells cultured toward Th17 phenotype or left as control (Th0). The data are presented as fold change over the expression level in Th0 control cells at 24 hours. The data show the average expressions and the SEM. Statistical significance of the results has been determined with the Student t test. (C) Representative CCR6 expression after 72 hours of culturing. (D) Schematic presentation of the microarray profiling showing the culturing conditions used and the sampling time points.