THE EPITOPE STABILITY OF GROUP-1 AND GROUP-2 ALLERGENS IN MITE EXTRACTS

Background: Commercial mite allergenic extracts sold in the US are pre pared with whole body mites in buffer solutions containing 50% glycero l. Mite extracts were reported to contain large number of proteolytic enzymes and their structural integrity in aqueous solutions have not b een evaluated. Objective: To evaluate the epitope stabilities of Group 1 and 2 allergens in two commercial mite extracts used by Center for Biologics Evaluation and Research (CBER) as reference extracts, E5-Dp (Dermatophagoides pteronyssinus) and E5-Df (D. farinae). Methods: Epit ope stability was determined by using monoclonal antibodies in a Sandw ich ELISA. Samples were stored at four different temperatures and the amounts of Der p 1, Der p 2, Der f 1, and Der f 2 were determined at d ifferent time intervals. The overall stability of mite extracts was ev aluated by immunoblot and competition ELISA. Results: The epitope stab ility of these allergens varies: Der f 1 was stable for at least 3 yea rs and Der f 2 for 1 year when stored at 4 degrees C; Der p 1 and 2 we re less stable. None of the Group 1 and 2 allergens remained intact wh en stored at 50 degrees C. Immunoblot and competition ELISA data also showed similar trend of degradation as compared with extracts stored a t 4 degrees C for same length of time. Conclusion: With the exception of Der f 1, the amount of detectable epitopes in Group 1 and 2 allerge ns reduce rapidly after 1 year, especially at elevated temperatures. T he changes in allergen composition were also observed by immunoblottin g and in relative potency by ELISA competition assay. These findings a re highly relavent to the users of CBER's mite extracts as standards.