Characterization and cloning of a major high molecular weight house dust mite allergen (Der f 15) for dogs

Although house dust mites (HDM(s)) are important elicitors of canine allerg y, the low molecular weight molecules defined as major allergens for humans do not appear to be major allergens for dogs. Western blotting of Dermatop hagoides farinae (D. farinae) extracts with sera from sensitized dogs showe d that the majority of animals had IgE antibodies specific for two proteins of apparent molecular weights of 98 and 109 kDa (98/109 kDa). The N-termin al sequences of these two proteins were identical, suggesting they were ver y closely related, and sequencing of internal peptides showed the protein(s ) to have homology with insect chitinases. A purified preparation of 98/109 kDa proteins elicited positive intradermal skin tests (IDST(s)) in a group of well-characterized atopic dogs sensitized to D. farinae, but not in nor mal dogs. A rabbit polyclonal antiserum raised against the purified protein s was used to immunoscreen a D. farinae cDNA library. The mature coding reg ion of the isolated chitinase cDNA predicts a protein of 63.2 kDa; sequence analysis and glycan detection blotting suggest that the molecule is extens ively O-glycosylated. Monoclonal antibodies made against the purified nativ e protein were used to localize the chitinase in sections of whole D. farin ae mites. The protein displayed an intracellular distribution in the proven triculus and intestine of the mite, suggesting that it has a digestive, rat her than a moulting-related, function. The high prevalence of IgE antibodie s to this antigen in canine atopic dermatitis makes it a major HDM allergen for dogs, and the protein has been formally designated Der f 15. (C) 2001 Elsevier Science B.V. All rights reserved.