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House dust mites produce well-known allergens for asthma and allergy among children. To study house dust mite allergen exposure level in northeast China and characterize its association with indoor environmental factors and cleaning habits, we inspected 399 homes in Tianjin area and collected dust from mattresses. Dermatophagoides farinae (Der f) and Dermatophagoides pteronyssinus (Der p) were detected by the enzyme-linked immunosorbent assay (ELISA) method. The medians of total allergen concentrations for spring, summer, autumn, and winter were 524 ng/g, 351 ng/g, 1022 ng/g, and 1010 ng/g. High indoor air relative humidity (RH), low air change rate, indoor dampness, and frequent changing of quilt cover/bedsheet/pillow case were significantly associated with high house dust mite allergen concentration (relative risk [RR]: RH, 1.18-1.34; air change rate, 0.97-1.00; dampness, 2.92-3.83; changing quilt cover/bedsheet/pillow case, 0.66-0.75). The decrease in the absolute humidity gradient between indoors and outdoors that occurs with increased air change rate may explain why a high ventilation reduces house dust mite allergen concentration. The findings of this study show the importance of ventilation and cleaning for controlling house dust mite allergens. We found that the decrease in additional absolute humidity (e.g., humidity (indoor)-humidity (outdoor)) with increased air change rate may be the main reason that a high ventilation rate reduces house dust mite allergen concentration. Ventilation and cleaning should be both considered for creating a healthy home environment.