title: Aptamer Lateral Flow Assays for Ultrasensitive Detection of β-Conglutin Combining Recombinase Polymerase Amplification and Tailed Primers.
abstract: In this work, different methodologies were evaluated in search of robust, simple, rapid, ultrasensitive, and user-friendly lateral flow aptamer assays. In one approach, we developed a competitive based lateral flow aptamer assay, in which β-conglutin immobilized on the test line of a nitrocellulose membrane and β-conglutin in the test sample compete for binding to AuNP labeled aptamer. The control line exploits an immobilized DNA probe complementary to the labeled aptamer, forcing displacement of the aptamer from the β-conglutin-aptamer complex. In a second approach, the competition for aptamer binding takes place off-strip, and following competition, aptamer bound to the immobilized β-conglutin is eluted and used as a template for isothermal recombinase polymerase amplification, exploiting tailed primers, resulting in an amplicon of a duplex flanked by single stranded DNA tails. The amplicon is rapidly and quantitatively detected using a nucleic acid lateral flow with an immobilized capture probe and a gold nanoparticle labeled reporter probe. The competitive lateral flow is completed in just 5 min, achieving a detection limit of 55 pM (1.1 fmol), and the combined competitive-amplification lateral flow requires just 30 min, with a detection limit of 9 fM (0.17 amol).
authors.affiliation: Nanobiotechnology and Bioanalysis group, Department of Chemical Engineering, Universitat Rovira I Virgili , 43007 Tarragona, Spain.
authors.forename: Miriam
authors.initials: M
authors.lastname: Jauset-Rubio
authors.affiliation: Nanobiotechnology and Bioanalysis group, Department of Chemical Engineering, Universitat Rovira I Virgili , 43007 Tarragona, Spain.
authors.forename: Markéta
authors.initials: M
authors.lastname: Svobodová
authors.affiliation: Nanobiotechnology and Bioanalysis group, Department of Chemical Engineering, Universitat Rovira I Virgili , 43007 Tarragona, Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: Mairal
authors.affiliation: Institute of Cellular Medicine, Diagnostic and Therapeutic Technologies Group, Newcastle University , Newcastle upon Tyne, NE2 4HH, U.K.
authors.forename: Calum
authors.initials: C
authors.lastname: McNeil
authors.affiliation: Institute of Cellular Medicine, Diagnostic and Therapeutic Technologies Group, Newcastle University , Newcastle upon Tyne, NE2 4HH, U.K.
authors.forename: Neil
authors.initials: N
authors.lastname: Keegan
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, Department of Chemical Engineering, Universitat Rovira I Virgili , 43007 Tarragona, Spain.|Institució Catalana de Recerca I Estudis Avancats , Passeig Lluís Companys 23, 08010 Barcelona, Spain.
authors.forename: Ciara K
authors.initials: CK
authors.lastname: O'Sullivan
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000235:genetics; D015374:Biosensing Techniques / Q000379:methods*; D015342:DNA Probes / Q000235:genetics; D006046:Gold / Q000737:chemistry; D057230:Limit of Detection; D031297:Lupinus; D053768:Metal Nanoparticles / Q000737:chemistry; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D055314:Seed Storage Proteins / Q000032:analysis* / Q000737:chemistry
chemical_list: D052157:Aptamers, Nucleotide; D015342:DNA Probes; D055314:Seed Storage Proteins; C042457:conglutin protein, Lupinus angustifolius; D006046:Gold