Gene expression profile of CD4+ T-cells restimulated by anti-CD3 and anti-ICOS compared to anti-CD3 alone

Engagement of the ICOS receptor represents a key event in a process that culminates in Bcl6 expression and acquisition of the TFH and TFR phenotype. To better understand the essentials of ICOS-mediated signaling pathway, we profiled the changes in gene expression elicited after co-ligation of ICOS and CD3 compared with CD3 ligation alone. Naïve CD4+ T-cells were purified from single cell suspensions of B6 spleen and stimulated with anti-CD3 and anti-CD28 for 2d followed by resting overnight before 20 min of incubation with anti-CD3 and/or anti-ICOS and cross-linking with goat anti-hamster Ab for 8h. RNA was prepared, amplified, labeled and hybridized to Mouse Gene 1.0 ST Array (Affymetrix), in quadruplicates for anti-CD3 ligation and duplicates for anti-CD3/ anti-ICOS co-ligation.