Two species of mites of the genus Dermatophagoides are common in house dust and make a major contribution to the allergen content of house dust. As judged by skin tests and the radioallergosorbent test, these mites, D. pteronyssinus and D. farinae, demonstrate extensive cross-reactivity. When the major allergen from D. pteronyssinus, antigen P1, was compared with the equivalent allergen from D. farinae, these two proteins were found to have both species-specific and common antigenic determinants. With specifically purified antibodies directed against the common determinants, we developed a radioimmunoassay for these antigen P1 equivalent proteins in mite extracts as well as house dust extracts. The quantity of allergen in dust from 63 houses (255 samples) was measured, and values ranged from less than 100 to greater than 100,000 ng/gm of fine dust. The correlation between antigen P1 equivalent (ng/gm) and the number of mites per gram of dust (identified by microscopy) was very good (r = 0.74; p less than 0.001). Furthermore, this correlation was not affected by the species of mite in the dust sample. Of 9243 mites identified, 95% were of the genus Dermatophagoides, and of these, 16.4% and 18% could be positively identified as D. farinae and D. pteronyssinus, respectively. When the assay for antigen P1 equivalent was applied to isolated components of mite cultures of both species, significant allergen was found in the mite bodies, cuticles, and their excreta (fecal particles). For the house-dust samples, less than 10% of the allergen measured could be explained by the allergen content of mite bodies. Our results demonstrate that a radioimmunoassay for antigen P1 equivalent can provide an accurate and simple means of assessing the quantity of mite-derived allergen in dust-mite extracts, house-dust extracts, or house dust.