T follicular helper (T FH) cells are central to the development and regulation of T cell-dependent humoral immune responses. The transcriptional repressor BCL6 is required for T FH responses, but the kinetics of BCL6 protein expression in activated CD4+ T cells have not been established. We measured BCL6 expression during T FH cell development at the single-cell level using intracellular staining and YFP-BCL6 fusion protein reporter mice. BCL6 was immediately upregulated in all dividing T cells during dendritic cell–T cell interactions. A second wave of early BCL6 expression coincided with the induction of CXCR5, resulting in a distinct and stable T FH cell population. Cognate B cells were not required for the induction of BCL6, but supported the expansion of T FH cells. These data suggest that BCL6 participates in very early events in T FH cell development, and that repeated encounters with APCs reinforce BCL6 expression, thereby establishing the T FH cell phenotype.