Background: Amino acid-based formulas (AAFs) are used for the dietary management of cowʼs milk allergy (CMA). Whether AAFs have the potential to prevent the development and/or symptoms of CMA is not known.Objective: The present study evaluated the preventive effects of an amino acid (AA)-based diet on allergic sensitization and symptoms of CMA in mice and aimed to provide insight into the underlying mechanism. Methods: C3H/HeOuJ mice were sensitized with whey protein or with phosphate-buffered saline as sham-sensitized control. Starting 2 weeks before sensitization, mice were fed with either a protein-based diet or an AA-based diet with an AA composition based on that of the AAF Neocate, a commercially available AAF prescribed for the dietary management of CMA. Upon challenge, allergic symptoms, mast cell degranulation, whey-specific immunoglobulin levels, and FoxP3 + cell counts in jejunum sections were assessed.Results: Compared to mice fed with the protein-based diet, AA-fed mice had significantly lower acute allergic skin responses. Moreover, the AA-based diet prevented the whey-induced symptoms of anaphylaxis and drop in body temperature. Whereas the AA-based diet had no effect on the levels of serum IgE and mucosal mast cell protease-1 (mMCP-1), AA-fed mice had significantly lower serum IgG2a levels and tended to have lower IgG1 levels (P = .076). In addition, the AA-based diet prevented the whey-induced decrease in FoxP3 + cells. In sham-sensitized mice, no differences between the two diets were observed in any of the tested parameters. Conclusion: This study demonstrates that an AA-based diet can at least partially prevent allergic symptoms of CMA in mice. Differences in FoxP3 + cell counts and serum levels of IgG2a and IgG1 may suggest enhanced antiinflammatory and tolerizing capacities in AA-fed mice. This, combined with ---This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.How to cite this article: van Sadelhoff JHJ, Hogenkamp A, Wiertsema SP, et al. A free amino acid-based diet partially prevents symptoms of cowʼs milk allergy in mice after oral sensitization with whey. Immun Inflamm Dis. 2020;8:93-105.