title: Detection of the cyanobacterial toxin, microcystin-LR, using a novel recombinant antibody-based optical-planar waveguide platform.
abstract: Microcystins are a major group of cyanobacterial heptapeptide toxins found in freshwater and brackish environments. There is currently an urgent requirement for highly-sensitive, rapid and in-expensive detection methodologies for these toxins. A novel single chain fragment variable (scFv) fragment was generated and is the first known report of a recombinant anti-microcystin avian antibody. In a surface plasmon resonance-based immunoassay, the antibody fragment displayed cross-reactivity with seven microcystin congeners (microcystin-leucine-arginine (MC-LR) 100%, microcystin-tyrosine-arginine (MC-YR) 79.7%, microcystin-leucine-alanine (MC-LA) 74.8%, microcystin-leucine-phenylalanine (MC-LF) 67.5%, microcystin-leucine-tryptophan (MC-LW) 63.7%, microcystin-arginine-arginine (MC-RR) 60.1% and nodularin (Nod) 69.3%, % cross reactivity). Following directed molecular evolution of the parental clone the resultant affinity-enhanced antibody fragment was applied in an optimized fluorescence immunoassay on a planar waveguide detection system. This novel immuno-sensing format can detect free microcystin-LR with a functional limit of detection of 0.19 ng mL(-1)and a detection range of 0.21-5.9 ng mL(-1). The assay is highly reproducible (displaying percentage coefficients of variance below 8% for intra-day assays and below 11% for inter-day assays), utilizes an inexpensive cartridge system with low reagent volumes and can be completed in less than twenty minutes.
authors.affiliation: School of Biotechnology, National Centre for Sensor Research and Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland. Electronic address: caroline.s.murphy@dcu.ie.
authors.forename: Caroline
authors.initials: C
authors.lastname: Murphy
authors.affiliation: School of Biotechnology, National Centre for Sensor Research and Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland.
authors.forename: Edwina
authors.initials: E
authors.lastname: Stack
authors.affiliation: School of Biotechnology, National Centre for Sensor Research and Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland.
authors.forename: Svetlana
authors.initials: S
authors.lastname: Krivelo
authors.affiliation: School of Biotechnology, National Centre for Sensor Research and Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland.
authors.forename: Daniel A
authors.initials: DA
authors.lastname: McPartlin
authors.affiliation: School of Biotechnology, National Centre for Sensor Research and Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland.
authors.forename: Barry
authors.initials: B
authors.lastname: Byrne
authors.affiliation: MBio Diagnostics Inc., Boulder, CO, 80301, USA.
authors.forename: Charles
authors.initials: C
authors.lastname: Greef
authors.affiliation: MBio Diagnostics Inc., Boulder, CO, 80301, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Lochhead
authors.affiliation: MBio Diagnostics Inc., Boulder, CO, 80301, USA.
authors.forename: Greg
authors.initials: G
authors.lastname: Husar
authors.affiliation: Institute for Global Food Security (IGFS), School of Biological Sciences, Queen's University, Malone Road, 18-30 Belfast, BT9 5BN, Northern Ireland, United Kingdom.
authors.forename: Shauna
authors.initials: S
authors.lastname: Devlin
authors.affiliation: Institute for Global Food Security (IGFS), School of Biological Sciences, Queen's University, Malone Road, 18-30 Belfast, BT9 5BN, Northern Ireland, United Kingdom.
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Elliott
authors.affiliation: School of Biotechnology, National Centre for Sensor Research and Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: O'Kennedy
mesh_terms: D015374:Biosensing Techniques; D000458:Cyanobacteria / Q000737:chemistry; D005618:Fresh Water / Q000032:analysis; D007118:Immunoassay; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000276:immunology; D008387:Marine Toxins; D052998:Microcystins / Q000737:chemistry / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology; D020349:Surface Plasmon Resonance
chemical_list: D007128:Immunoglobulin Fragments; D008387:Marine Toxins; D052998:Microcystins; D011994:Recombinant Proteins; C057862:cyanoginosin LR
keywords: Cyanobacteria; Fluorescent planar wave-guide; Immunoassay; Microcystin; Recombinant Antibody