Abstract: Wheat is the most widely produced and consumed food in the world, and it is also one of the eight allergenic foods recognized in the world. Gliadin (Glia) is one of the main allergens that induce wheat allergy. This study aimed to explore the effects of high intensity water bath ultrasound (HIWU) on the structure and antigenicity of Glia. Glia was treated with different ultrasonic power. The results of Western blotting showed that there was a strong immune response before and after ultrasound treatment. Non-competitive enzyme-linked immunosorbent assay (ELISA) showed that HIWU significantly reduced the antigenicity of Glia, and the highest immunosuppression rate was 33.03%±1.72% after 480 W HIWU. Furthermore, SDS-PAGE showed that HIWU did not produce small molecular subunits of Glia but cause aggregation of some protein subunits. HIWU reduced the content of α-helix and β-sheet of Glia to make it disordered. HIWU caused quenching of the fluorescence of Glia, and the content of free sulfhydryl groups in protein increased. Meanwhile, Raman spectroscopy analysis showed that the main conformation of the disulfide bond changed from g-g-g to g-g-t, tyrosine and tryptophan changed from the original embedded state to the exposed state. HIWU altered the secondary and tertiary structures of Glia, resulting in the deletion or destruction of Glia epitopes, thereby effectively inhibiting the antigenicity of Glia. This study will broaden the application of ultrasonic technology in cereal products and provide a theoretical reference for the production of hypoallergenic wheat food.