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We examined the anti-allergic effect of strawberry extract on human peripheral blood mononuclear cells (PBMCs) and atopic dermatitis model mice NC/NgaTndCrlj. The addition of strawberry extract suppressed total IgE production in the cedar pollen antigen Cry j 1-stimulated PBMCs. Flow cytometric analysis showed that strawberry extract decreased the rate of CD3+CD4+ helper T cells by 17.3% and increased the rate of CD3+CD8+ cytotoxic T cells by 19.7% in PBMCs. Moreover, the extract inhibited the expression level of GATA3 that is the master regulator of type 2 helper T cells (Th2) in human primary pan T cells isolated from PBMCs. Oral administration of strawberry extract lowered dermatitis scores and serum IgE levels in mice. In addition, it also decreased the GATA3 expression level in mouse blood cells. These results revealed that strawberry extract suppressed the severity of atopic dermatitis through the down-regulation of serum IgE by inhibition of Th2 differentiation.