Non-protease native allergens partially purified from bodies of eight domestic mites using p-aminobenzamidine ligand

• T. Erban ^[1] ; R. Klubal ^[2]
  1. [1] Crop Research Institute

     Crop Research Institute

     Chequia

     
  2. [2] Heritage Valley Health System

     Heritage Valley Health System

     Borough of Beaver, Estados Unidos

     
• Localización: Allergologia et immunopathologia: International journal for clinical and investigate allergology and clinical immunology, ISSN-e 1578-1267, ISSN 0301-0546, Vol. 46, Nº. 3, 2018, págs. 218-225
• Idioma: inglés
• Texto completo no disponible (Saber más ...)
• Resumen

  • Background Optimised purification steps for concentrating trace target native antigens are needed. Combining the p-aminobenzamidine ligand with protease inactivation enables partial purification of mite non-protease allergens lacking proteases.

    Objective We sought to analyse in detail proteins obtained using this method from eight species of synanthropic acaridid mites and tested IgE reactivity using pooled human sera.

    Materials and methods Proteins affinity bound to p-aminobenzamidine as a ligand were identified by MALDI TOF/TOF. After electroblotting, the proteins were visualised using the fluorescent SYPRO-Ruby protein blot stain, and IgE reactivity was further analysed using pooled human sera collected from patients allergic to house dust mites.

    Results MS/MS identification confirmed previous results that no proteases were purified. Protein patterns corresponding to the allergens Der f 7, Der f 30 and actins indicated that these proteins are purified using p-aminobenzamidine and are present across a wide spectrum of acaridid mites. When using Dermatophagoides farinae, apolipophorins (Der f 14), chitinase-like Der f 15 and 18, 70-kDa heat shock protein, and a Der f Alt a10 allergen homolog (gi|37958173) were also detected. The target antigens tropomyosins and paramyosins showed similar IgE binding among the mite species tested. IgE reactivity with miscellaneous D. farinae antigen was also observed.

    Conclusions Partial purification of mite non-protease antigens using a strategy combining p-aminobenzamidine with protease inactivation was verified by 1D-E and 2D-E analyses. IgE binding to p-aminobenzamidine-purified native non-protease mite antigens was tested using pooled sera. This preliminary study allows for further work on individual serum samples, allowing confirmation of immunoreactivity.