Atopy patch tests (APTs) are considered a useful addition to standard testing procedures for the identification of allergens triggering the appearance of eczema in patients with atopic dermatitis (AD). The aim of the study was to evaluate the reproducibility of APTs with house dust mites by simultaneously patch testing AD patients. 85 subjects affected by AD, 36 males and 48 females, aged from 1 to 52 years, were patch tested with Dermatophagoides mix (D. pteron/D. farinae) 20% pet. In our patients, the percentage of agreement between the frequency of positive and negative responses to Dermatophagoides mix was 96.7%. Reproducibility was highly satisfactory, as evaluated by k-value (0.953). As regards the agreement in the intensity of APT responses to Dermatophagoides mix between the right and left sides, we found that reproducibility was also highly satisfactory (k = 0.851). In conclusion, for Dermatophagoides APTs, the agreement rate was similar to that of standard patch tests, and therefore APTs may be considered sufficiently reproducible to be employed as a diagnostic testing procedure.