title: Enzyme-free amplification and detection of bovine viral diarrhea virus RNA using hybridization chain reaction and gold nanoparticles.
abstract: A novel bovine viral diarrheal virus (BVDV)-RNA detection method was developed using a combination of the amplification capability of hybridization chain reaction (HCR) with the sensitivity of an unmodified-gold nanoparticle (AuNP) colorimetric detection assay. Two auxiliary probes were designed to target a conserved RNA sequence among the BVDV isolates. The complementary target BVDV-RNA was used as the initiator to trigger a cascade of hybridization events to yield nicked double-helix DNA analogous to the alternating copolymers. DNA in the form of a nicked double helix did not prevent salt-induced aggregation of AuNPs. In contrast, in the absence of the complementary target BVDV-RNA, free hairpins with single-stranded sticky ends adsorbed onto the AuNPs, stabilize them, and prevent salt-induced aggregation of the AuNP. The limit of detection (LOD) for the BVDV-RNA was estimated to be 0.008 tissue culture infective dose (TCID50)/reaction. The method developed was highly selective and specific to detect BVDV isolates in clinical samples. This protocol offers a rapid, simple, and cost-effective assay for detecting BVDV.
authors.affiliation: Department of Biology, Faculty of Science, Shahid Chamran University of Ahvaz, Ahvaz, 6135743135, Iran.
authors.forename: Shahrzad
authors.initials: S
authors.lastname: Ghasemi Monjezi
authors.affiliation: Department of Biology, Faculty of Science, Shahid Chamran University of Ahvaz, Ahvaz, 6135743135, Iran. e.tofighi@yahoo.com.
authors.forename: Seyedeh Elham
authors.initials: SE
authors.lastname: Rezatofighi
authors.affiliation: Department of animal science and food, Khouzastan Ramin Agriculture and Natural Resources University, Ahvaz, Iran.
authors.forename: Khalil
authors.initials: K
authors.lastname: Mirzadeh
authors.affiliation: Department of Chemistry, Faculty of Science, Shahid Chamran University of Ahvaz, Ahvaz, Iran.
authors.forename: Saadat
authors.initials: S
authors.lastname: Rastegarzadeh
mesh_terms: D003124:Colorimetry / Q000379:methods*; D001908:Diarrhea Viruses, Bovine Viral / Q000235:genetics / Q000302:isolation & purification*; D006046:Gold; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D053758:Nanoparticles; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012367:RNA, Viral / Q000032:analysis* / Q000235:genetics; D012680:Sensitivity and Specificity
chemical_list: D012367:RNA, Viral; D006046:Gold
keywords: Bovine viral diarrhea virus; Detection; Gold nanoparticles; Hybridization chain reaction