Background: Little is known about the role of molecular diagnosis in house dust mite (HDM) allergy. Objective: In this study, we investigated the association between the sensitization profile of adolescent and adult HDM-allergic patients and asthma in a region with high rates of exposure to HDM. Methods: We conducted a cross-sectional study of 384 HDM-allergic patients (38.5%, males; median age, 28 years). A total of 368 patients (95.8%) had rhinitis, and 175 (45.6%) had asthma. Specific IgE (sIgE) to Dermatophagoides pteronyssinus, nDer p 1, rDer p 2, and rPen a 1 was measured in all patients. sIgE to Lepidoglyphus destructor was measured in patients (n=301) with a positive skin test result. Results: Significantly higher concentrations of sIgE to Der p 1 and sIgE to Der p 2 were observed in patients with asthma than in those without asthma. The proportion of asthmatic patients was higher among individuals who reacted (≥0.35 kU A /L) to both Der p 1 and Der p 2 (147/291, 50.5%) than among those who reacted to only 1 allergen (either Der p 1 or Der p 2, 18/55, 32.7%) or neither allergen (10/38, 26.3%, P=.002). Reactivity to both allergens was associated with asthma after adjusting for age and sex (OR, 2.87; 95%CI,. Higher concentrations of sIgE to L destructor were observed in patients with asthma than in patients without asthma. Tropomyosin sIgE ≥0.35 kU A /L was detected in only 6 individuals (1.6%). Conclusions: L destructor may be a relevant allergen in high-exposure areas. Dual sensitization (ie, IgE to both Der p 1 and Der p 2) may help to identify HDM-allergic patients who are at risk of asthma.