The antigenic analysis of Dermatophagoides pteronyssinus extract was performed by using antiserum obtained in rabbits by immunization with mite extract in Freund's adjuvant. Immunodiffusion and immunoelectrophoretic analysis revealed the presence of many constituents, most of which were common to human dander, house dust and Dermatophagoides farinae extracts. By fractionation of the mite extract on Sephadex G-75 and DEAE-Sephadex it was shown that the highest allergenic activity was related to a family of heat-stable molecules weighing about 30,000. The antigenic analysis of chromatographic most active fraction revealed the presence of three heat-stable constituents, one of which is not shared by nutrient medium, or by D. farinae extract. The fractionation of D. pteronyssinus extract by preparative isoelectrophocusing on sucrose gradient (pH range 3-10) showed that the two fractions (pH 3.5 and pH 5.0) containing the major allergenic activity (measured by the RAST-inhibition capacity) were also able to induce a significant in vitro activation of lymphocytes from allergic patients. When the anti-mite rabbit gammaglobulins were fixed to an immunosorbent and incubated with a house dust extract, it was constantly possible to induce a strong decrease or a complete loss of allergenicity of the dust extract in man. This is in favour of an allergenic identity between dust and mite.