title: Magnetic Bead/Gold Nanoparticle Double-Labeled Primers for Electrochemical Detection of Isothermal Amplified Leishmania DNA.
abstract: A novel methodology for the isothermal amplification of Leishmania DNA using labeled primers combined with the advantages of magnetic purification/preconcentration and the use of gold nanoparticle (AuNP) tags for the sensitive electrochemical detection of such amplified DNA is developed. Primers labeled with AuNPs and magnetic beads (MBs) are used for the first time for the isothermal amplification reaction, being the amplified product ready for the electrochemical detection. The electrocatalytic activity of the AuNP tags toward the hydrogen evolution reaction allows the rapid quantification of the DNA on screen-printed carbon electrodes. Amplified products from the blood of dogs with Leishmania (positive samples) are discriminated from those of healthy dogs (blank samples). Quantitative studies demonstrate that the optimized method allows us to detect less than one parasite per microliter of blood (8 × 10(-3) parasites in the isothermal amplification reaction). This pioneering approach is much more sensitive than traditional methods based on real-time polymerase chain reaction (PCR), and is also more rapid, cheap, and user-friendly.
authors.affiliation: Catalan Institute of Nanoscience and Nanotechnology (ICN2), CSIC and Barcelona Institute of Science and Technology, Campus UAB, 08193, Barcelona, Spain.
authors.forename: Alfredo
authors.initials: A
authors.lastname: de la Escosura-Muñiz
authors.affiliation: Catalan Institute of Nanoscience and Nanotechnology (ICN2), CSIC and Barcelona Institute of Science and Technology, Campus UAB, 08193, Barcelona, Spain.
authors.forename: Luis
authors.initials: L
authors.lastname: Baptista-Pires
authors.affiliation: Vetgenomics, Edifici Eureka, Parc de Recerca UAB, 08193, Bellaterra (Barcelona), Spain.
authors.forename: Lorena
authors.initials: L
authors.lastname: Serrano
authors.affiliation: Vetgenomics, Edifici Eureka, Parc de Recerca UAB, 08193, Bellaterra (Barcelona), Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Altet
authors.affiliation: Vetgenomics, Edifici Eureka, Parc de Recerca UAB, 08193, Bellaterra (Barcelona), Spain.|Department of Animal and Food Science, Autonomous University of Barcelona, Campus UAB, 08193, Bellaterra (Barcelona), Spain.
authors.forename: Olga
authors.initials: O
authors.lastname: Francino
authors.affiliation: Department of Animal and Food Science, Autonomous University of Barcelona, Campus UAB, 08193, Bellaterra (Barcelona), Spain.|Centre for Research in Agricultural Genomics-CSIC-IRTA-UAB-UB, Campus UAB, 08193, Cerdanyola del Valles, Catalonia, Spain.
authors.forename: Armand
authors.initials: A
authors.lastname: Sánchez
authors.affiliation: Catalan Institute of Nanoscience and Nanotechnology (ICN2), CSIC and Barcelona Institute of Science and Technology, Campus UAB, 08193, Barcelona, Spain.|ICREA-Institucio Catalana de Recerca i Estudis Avançats, 08010, Barcelona, Spain.
authors.forename: Arben
authors.initials: A
authors.lastname: Merkoçi
mesh_terms: D000818:Animals; D001681:Biological Assay; D004247:DNA / Q000032:analysis* / Q000235:genetics; D017931:DNA Primers / Q000378:metabolism*; D004563:Electrochemistry / Q000379:methods*; D004587:Electrophoresis, Agar Gel; D006046:Gold / Q000737:chemistry*; D007891:Leishmania / Q000235:genetics*; D008280:Magnetics; D053768:Metal Nanoparticles / Q000737:chemistry*; D008863:Microspheres; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D010271:Parasites / Q000235:genetics; D013194:Staining and Labeling
chemical_list: D017931:DNA Primers; D006046:Gold; D004247:DNA
keywords: Leishmania; electrocatalytic detection; gold nanoparticles; isothermal DNA amplification; magnetic beads