This study aimed to evaluate the anti-atopic dermatitis (AD) effects of Isatidis folium water extract (WIF) using a 2,4-dinitrochlorobenzene (DNCB)-induced AD-like mouse model and to investigate the underlying mechanism using tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ)-activated HaCaT cells. The gene discussed is IFNG; the disease is atopic eczema.