title: Ultrasensitive and rapid detection of β-conglutin combining aptamers and isothermal recombinase polymerase amplification.
abstract: Lupin is increasingly being used in a variety of food products due to its nutritional, functional and nutraceutical properties. However, several examples of severe and even fatal food-associated anaphylaxis due to lupin inhalation or ingestion have been reported, resulting in the lupin subunit β-conglutin, being defined as the Lup an 1 allergen by the International Union of Immunological Societies (IUIS) in 2008. Here, we report an innovative method termed aptamer-recombinase polymerase amplification (Apta-RPA) exploiting the affinity and specificity of a DNA aptamer selected against the anaphylactic β-conglutin allergen termed β-conglutin binding aptamer II (β-CBA II), facilitating ultrasensitive detection via isothermal amplification. Combining magnetic beads as the solid phase with Apta-RPA detection, the total assay time was reduced from 210 min to just 25 min, with a limit of detection of 3.5 × 10-11M, demonstrating a rapid and ultrasensitive generic methodology that can be used with any aptamer. Future work will focus on further simplification of the assay to a lateral flow format. Graphical Abstract Schematic representation of the rapid and novel bead-based Apta-RPA assay.
authors.affiliation: Nanobiotechnology and Bioanalysis Group, INTERFIBIO Consolidated Group, Department of Chemical Engineering, Universitat Rovira I Virgili, Campus Sescelades, Avinguda dels Països Catalans, 26, 43007, Tarragona, Spain.
authors.forename: Miriam
authors.initials: M
authors.lastname: Jauset-Rubio
authors.affiliation: Nanobiotechnology and Bioanalysis Group, INTERFIBIO Consolidated Group, Department of Chemical Engineering, Universitat Rovira I Virgili, Campus Sescelades, Avinguda dels Països Catalans, 26, 43007, Tarragona, Spain.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Sabaté Del Río
authors.affiliation: Nanobiotechnology and Bioanalysis Group, INTERFIBIO Consolidated Group, Department of Chemical Engineering, Universitat Rovira I Virgili, Campus Sescelades, Avinguda dels Països Catalans, 26, 43007, Tarragona, Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: Mairal
authors.affiliation: Nanobiotechnology and Bioanalysis Group, INTERFIBIO Consolidated Group, Department of Chemical Engineering, Universitat Rovira I Virgili, Campus Sescelades, Avinguda dels Països Catalans, 26, 43007, Tarragona, Spain.
authors.forename: Markéta
authors.initials: M
authors.lastname: Svobodová
authors.affiliation: Department of Chemistry, Faculty of Science, King Abdulaziz University, P. O. Box 80203, Jeddah, 21589, Kingdom of Saudi Arabia.
authors.forename: Mohammad S
authors.initials: MS
authors.lastname: El-Shahawi
authors.affiliation: Department of Chemistry, Faculty of Science, King Abdulaziz University, P. O. Box 80203, Jeddah, 21589, Kingdom of Saudi Arabia.
authors.forename: Abdulaziz S
authors.initials: AS
authors.lastname: Bashammakh
authors.affiliation: Department of Chemistry, Faculty of Science, King Abdulaziz University, P. O. Box 80203, Jeddah, 21589, Kingdom of Saudi Arabia.
authors.forename: Abdulrahman O
authors.initials: AO
authors.lastname: Alyoubi
authors.affiliation: Nanobiotechnology and Bioanalysis Group, INTERFIBIO Consolidated Group, Department of Chemical Engineering, Universitat Rovira I Virgili, Campus Sescelades, Avinguda dels Països Catalans, 26, 43007, Tarragona, Spain. ciara.osullivan@urv.cat.|Institució Catalana de Recerca I Estudis Avancats, Passeig Lluís Companys 23, 08010, Barcelona, Spain. ciara.osullivan@urv.cat.
authors.forename: Ciara K
authors.initials: CK
authors.lastname: O'Sullivan
mesh_terms: D000485:Allergens / Q000032:analysis*; D052157:Aptamers, Nucleotide / Q000737:chemistry*; D015374:Biosensing Techniques / Q000379:methods*; D031297:Lupinus / Q000737:chemistry*; D016133:Polymerase Chain Reaction / Q000379:methods; D045522:Recombinases / Q000737:chemistry; D052156:SELEX Aptamer Technique / Q000379:methods; D055314:Seed Storage Proteins / Q000032:analysis*
chemical_list: D000485:Allergens; D052157:Aptamers, Nucleotide; D045522:Recombinases; D055314:Seed Storage Proteins; C042457:conglutin protein, Lupinus angustifolius
keywords: Apta-PCR; Apta-RPA; Aptamer; Lupin; Recombinase polymerase amplification