High throughput screening: a rapid way to recombinant allergens.
Complex allergenic sources such as moulds, foods and mites contain large panels of IgE-binding molecules which need to be cloned, produced and characterized in order to mimic the entire allergenicity of whole extracts reconstituted by mixing single standardized recombinant allergens. Phage display of cDNA libraries allows selective enrichment of allergen-expressing clones using IgE from allergic patients. For the characterization of all different clones present in enriched cDNA libraries in a fast and cost-effective way, however, high-throughput screening technology is required. We have used a high-throughput, quantitative technology for fast identification of all different clones present in selectively enriched phage surface-displayed cDNA libraries in order to characterize whole allergenic repertoires from complex allergenic sources. The strategy, based on a combination of phage display and high-density arrays, allowed fast discovery of panels of related structures from different allergenic sources. They cover secreted, cytoplasmic and structural proteins with or without enzymatic activity and offer a rational explanation for the IgE-mediated cross-reactivity frequently encountered in clinical practice.
Crameri
R
R
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland.
D000485:Allergens / Q000235:genetics* / Q000627:therapeutic use; D000818:Animals; D003001:Cloning, Molecular / Q000379:methods*; D020650:Combinatorial Chemistry Techniques / Q000379:methods; D004274:DNA, Recombinant / Q000276:immunology* / Q000627:therapeutic use; D011994:Recombinant Proteins / Q000627:therapeutic use
D000485:Allergens; D004274:DNA, Recombinant; D011994:Recombinant Proteins