Wheat-dependent exercise-induced anaphylaxis (WDEIA) is characterized by anaphylactic reactions after wheat ingestion and physical exercise. IgE antibodies to recombinant ω5-gliadin are detectable in a majority of WDEIA patients, but other wheat allergens may also play a role in elicitation of WDEIA. Here, we performed a comprehensive analysis of IgE reactivity to different wheat proteins in 17 patients with confirmed WDEIA by ImmunoCAP research prototypes and a semi-quantitative microarray immunoassay with α/β/γ-gliadin, high-molecular-weight (HMW) glutenin, alpha-amylase inhibitor (AAI) dimer, and wheat lipid transfer protein (LTP). By ImmunoCAP, IgE to recombinant ω5-gliadin was detectable in 14/17 patients (82%), to α/β/γ-gliadin in 82% including the three patients lacking IgE to ω5-gliadin, and to HMW glutenin in 59%. The microarray revealed specifically γ-gliadin as the second most important allergen. These results demonstrate the additional diagnostic value of α/β- and γ-gliadin in particular in ω5-gliadin-negative patients in the diagnosis of WDEIA.