The relative contribution of T cell receptor‐versus CD28‐mediated signals in co‐stimulation of resting CD4 T cells is thought to influence their functional differentiation towards T helper (Th) 1 versus Th2 subsets. We have used a conventional and a mitogenic CD28‐specific monoclonal antibody to assess the effect of polyclonal T cell activation through CD28 alone on CD4 subset differentiation. In vivo, mitogenic but not conventional anti‐CD28 induces massive lymphocytosis, the Th2 cytokines interleukin (IL)‐4 and IL‐10, and Th2‐dependent immunoglobulin isotypes, most notably IgE. In vitro, it is shown that mitogenic anti‐CD28 primes for IL‐4‐dependent induction of IL‐4 expression much more efficiently than conventional co‐stimulation. At the molecular level, we show for the first time that the activation of the “Th2 promoting” transcription factor GATA‐3 requires co‐stimulation by CD28 and is also induced by mitogenic anti‐CD28 alone. We suggest that CD28‐dependent induction of GATA‐3 in concert with other transcription factors, which are preferentially induced by strong CD28‐signals, primes CD4 T cells for IL‐4‐dependent Th2 differentiaton.