title: Electrochemical Biosensors Combined with Isothermal Amplification for Quantitative Detection of Nucleic Acids.
abstract: In recent years, various isothermal amplification techniques have been developed as alternatives to polymerase chain reaction (PCR). The integration of isothermal amplification with electrical or electrochemical devices has enabled high-throughput nucleic acid-based assays with high sensitivity. We performed solid-phase rolling circle amplification (RCA) on the surface of a Au electrode, and detected RCA products in situ using chronocoulometry (CC) with [Ru (NH3)6]3+as the signaling molecule. Detection sensitivity for DNA and a microRNA (miR-143) was 100 fM and 1 pM, respectively. Furthermore, we conducted potentiometric DNA detection using an ethidium ion (Et+)-selective electrode (Et+ISE) for real-time monitoring of isothermal DNA amplification by primer-generation RCA (PG-RCA). The Et+ISE potential enabled real-time monitoring of the PG-RCA reaction in the range of 10 nM-1 μM of initial target DNA. Devices based on these electrochemical techniques represent a new strategy for replacing conventional PCR for on-site detection of nucleic acids of viruses or microorganisms.
authors.affiliation: Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Miyuki
authors.initials: M
authors.lastname: Tabata
authors.affiliation: Department of Chemistry, Zhejiang University, Hangzhou, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Yao
authors.affiliation: Department of Applied Chemistry, Graduate School of Science and Engineering, Keio University, Yokohama, Japan.
authors.forename: Ayaka
authors.initials: A
authors.lastname: Seichi
authors.affiliation: Department of Applied Chemistry, Graduate School of Science and Engineering, Keio University, Yokohama, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Suzuki
authors.affiliation: Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Tokyo, Japan. miyahara.bsr@tmd.ac.jp.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Miyahara
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods; D004247:DNA / Q000032:analysis / Q000235:genetics; D055664:Electrochemical Techniques / Q000295:instrumentation / Q000379:methods; D004566:Electrodes; D006046:Gold; D035683:MicroRNAs; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation / Q000379:methods; D016133:Polymerase Chain Reaction; D013696:Temperature
chemical_list: D035683:MicroRNAs; D006046:Gold; D004247:DNA
keywords: Biosensor; Chronocoulometry; Ion selective electrode; Isothermal nucleic acid amplification; PCR; microRNA