Currently no validated animal model is predictive of human responses in ranking purified dietary proteins in the prevalence or potency of food allergy in humans. Since the gastrointestinal (GI) microbiota is thought to influence oral tolerance, we hypothesize that a germ-free mouse model will more accurately predict atopic human responses than conventional mice. Germ-free C3H/HeN mice were immunized with 60 μg Ara h 2, BLG, or LOX by three weekly intraperitoneal (IP) injections with alum adjuvant. One week following the final immunization an IP challenge of 500 μg of Ara h 2, BLG, or LOX was administered. Thirty minutes post-challenge clinical scores were graded and body temperatures recorded. The presence of protein-specific IgE and mast cell protease concentrations in mouse sera were determined using ELISA. Upon challenge germ-free mice sensitized with Ara h 2 and BLG exhibited significantly more severe clinical scores (average 4) compared to germ-free mice immunized with LOX (average 1). Hypothermic responses in challenged mice differed between the three proteins post-challenge. Results indicate that this model can differentiate between potent and non-allergens based on temperature drop, clinical scores, and biomarkers. Additional proteins with known human exposure and allergenicity are needed to confirm the predictive accuracy.