The cDNA coding for one of the major allergens of Dermatophagoides pteronyssinus, Der p II, has been cloned and expressed in Escherichia coli as a fusion protein with glutathione S-transferase. The recombinant material (r Der p II) was tested in the skin of house-dust allergic patients, and was found to be almost as active as the purified native allergen (n Der p II) in stimulating immediate hypersensitivity reactions. The positivity to r Der p II in these patients, who were selected for their skin-test reactivity to house-dust extract (HDE) and whole mite extract (WME), was over 60%, and 80% of their sera contained detectable IgE antibody against the recombinant allergen. The level of specific IgE antibody against r Der p II was a mean of 40% of that against WME in the sera of patients who were skin test positive to both allergen preparations. In contrast to the house-dust allergic patients, none of a group of allergic patients whose clinical history did not implicate house dust, and who were skin test negative to HDE or WME, were positive to r Der p II. Of interest was the observation that only 37% of a group of subjects who were skin test positive to HDE and WME, but who reported no clinical history of allergic disease upon natural exposure to house dust, reacted to r Der p II. These results demonstrate the biological activity of r Der p II, and indicate, in persons skin test positive to house-dust mite extract, an association between reactivity to Der p II and the manifestation of clinical symptoms.