title: A magnetic nanobead-based bioassay provides sensitive detection of single- and biplex bacterial DNA using a portable AC susceptometer.
abstract: Bioassays relying on magnetic read-out using probe-tagged magnetic nanobeads are potential platforms for low-cost biodiagnostic devices for pathogen detection. For optimal assay performance it is crucial to apply an easy, efficient and robust bead-probe conjugation protocol. In this paper, sensitive (1.5 pM) singleplex detection of bacterial DNA sequences is demonstrated in a portable AC susceptometer by a magnetic nanobead-based bioassay principle; the volume-amplified magnetic nanobead detection assay (VAM-NDA). Two bead sizes, 100 and 250 nm, are investigated along with a highly efficient, rapid, robust, and stable conjugation chemistry relying on the avidin-biotin interaction for bead-probe attachment. Avidin-biotin conjugation gives easy control of the number of detection probes per bead; thus allowing for systematic investigation of the impact of varying the detection probe surface coverage upon bead immobilization in rolling circle amplified DNA-coils. The existence of an optimal surface coverage is discussed. Biplex VAM-NDA detection is for the first time demonstrated in the susceptometer: Semi-quantitative results are obtained and it is concluded that the concentration of DNA-coils in the incubation volume is of crucial importance for target quantification. The present findings bring the development of commercial biodiagnostic devices relying on the VAM-NDA further towards implementation in point-of-care and outpatient settings.
authors.affiliation: Department of Engineering Sciences, Division of Solid State Physics, Uppsala University, Uppsala, Sweden. mattias.stromberg@angstrom.uu.se.
authors.forename: Mattias
authors.initials: M
authors.lastname: Strömberg
authors.forename: Teresa
authors.initials: T
authors.lastname: Zardán Gómez de la Torre
authors.forename: Mats
authors.initials: M
authors.lastname: Nilsson
authors.forename: Peter
authors.initials: P
authors.lastname: Svedlindh
authors.forename: Maria
authors.initials: M
authors.lastname: Strømme
mesh_terms: D001360:Avidin / Q000737:chemistry; D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods*; D001710:Biotin / Q000737:chemistry; D004269:DNA, Bacterial / Q000032:analysis / Q000737:chemistry*; D004270:DNA, Circular / Q000737:chemistry; D004277:DNA, Single-Stranded / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics; D058185:Magnetite Nanoparticles / Q000032:analysis* / Q000737:chemistry; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation; D015345:Oligonucleotide Probes / Q000737:chemistry / Q000235:genetics; D014734:Vibrio cholerae / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D004270:DNA, Circular; D004277:DNA, Single-Stranded; D058185:Magnetite Nanoparticles; D015345:Oligonucleotide Probes; D001360:Avidin; D001710:Biotin
keywords: AC susceptometer; Brownian relaxation; Padlock probes; Probe-tagged magnetic beads; Rolling circle amplification