ABSTRACT
A novel method of catalytic hairpin assembly (CHA) induced dual signal enhancement is developed for rapid detection of miRNA based on fluorescence light-up silver nanocluster (Ag NC). By the hybridization of a hairpin DNA and a single-stranded DNA, a unique probe is firstly designed. In the terminals of this probe, DNA-Ag NCs can be formed and display very weak fluorescence. In the presence of the target miRNA, the reaction of CHA can be triggered, forming two kinds of double-stranded complexes, in which the terminal DNA-Ag NCs are in close proximity to G-rich overhangs and the fluorescent signal can be dramatically enhanced. Compared with many other enzyme-based amplification strategies, this one exhibits distinct advantages of simplicity in experimental operation and a rapid detection process (within 1â¯h). Moreover, this assay exhibits an excellent selectivity and is successfully applied in the detection of miRNAs in complex biological media, which confirms the reliability and practicality of this protocol.
Subject(s)
DNA Probes/chemistry , DNA/chemistry , Fluorescence , Light , Metal Nanoparticles/chemistry , MicroRNAs/analysis , Silver/chemistry , Catalysis , DNA Probes/chemical synthesis , Humans , Spectrometry, Fluorescence , Tumor Cells, CulturedABSTRACT
A sensitive surface-enhanced Raman scattering (SERS) approach has been developed for detection of microRNA (miRNA) based on target-triggered dual signal amplification including strand displancement amplification (SDA) and hybridization chain reaction (HCR). With the assistant of polymerase and nicking endonuclease (NEase), target miRNA combines with the single stranded template DNA to generate a great amount of trigger DNA which can induce HCR. Coupled the dual cycle amplification of SDA and HCR with the dual enhancement of gold nanoparticles (AuNPs), a low detection limit of 0.5â¯fM for miRNA is obtained using the proposed strategy. With high sensitivity, universality, rapid analysis, and high selectivity, this method has a great potential for detecting biomolecules with trace amounts in bioanalysis and clinical biomedicine.
Subject(s)
MicroRNAs/analysis , Spectrum Analysis, Raman/methods , Gold/chemistry , Limit of Detection , Metal Nanoparticles/chemistryABSTRACT
A novel and sensitive surface-enhanced Raman scattering (SERS) method is proposed for the assay of DNA methyltransferase (MTase) activity and evaluation of inhibitors by developing a target triggering primer generation-based multiple signal amplification strategy. By using of a duplex substrate for Dam MTase, two hairpin templates and a Raman probe, multiple signal amplification mode is achieved. Once recognized by Dam MTase, the duplex substrate can be cleaved by Dpn I endonuclease and two primers are released for triggering the multiple signal amplification reaction. Consequently, a wide dynamic range and remarkably high sensitivity are obtained under isothermal conditions. The detection limit is 2.57×10(-4)UmL(-1). This assay exhibits an excellent selectivity and is successfully applied in the screening of inhibitors for Dam MTase. In addition, this novel sensing system is potentially universal as the recognition element can be conveniently designed for other target analytes by changing the substrate of DNA MTase.
