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The development of facile and sensitive miRNA quantitative detection methods is a central challenge for the early diagnosis of miRNA-related diseases. Herein, we propose a strategy for a liposome-encoded magnetic bead-based DNA toehold-mediated DNA circuit for the simple and sensitive detection of miRNA based on a toehold-mediated circular strand displacement reaction (TCSDR) coupled with a personal glucometer (PGM ). In this strategy, a glucoamylase-encapsulated liposomes (GELs)-encoded magnetic bead (GELs-MB) probe is designed to integrate target binding, magnetic separation, and signal response. Upon sensing the target miRNA-21, a GELs-MB probe-based toehold-mediated circular strand displacement reaction (TCSDR) was initiated with the help of fuel-DNA, constructing a DNA circuit system, and realizing target recycling amplification and the disassembly of the liposomes. The disassembled liposomes were finally removed via magnetic separation, and the encapsulated glucoamylase was liberated to catalyze amylose hydrolysis with multiple turnovers to glucose for a PGM readout. Benefiting from target recycling amplification initiated by the toehold-mediated DNA circuit and the liposome multiple-label amplification, a small quantity of target miRNA-21 can be transformed into a large glucose signal. The strategy realized the quantification of miRNA-21 down to a level of 0.7 fM without enzymatic amplification or precise instrumentation. Moreover, the high-density GELs-MB probe allows the sensitive detection of miRNA-21 to be accomplished within 1.5 h. Furthermore, this strategy exhibits the advantages of specificity and simplicity, since a toehold-mediated strand displacement reaction, magnetic separation and portable PGM were used. Importantly, this strategy has been demonstrated to allow the high-confidence quantification of miRNA. Therefore, with the advantages of low cost, ease of use, portability, and sensitivity, the reported method holds great potential for the early diagnosis of miRNA-related diseases.

Development of reliable and affordable quantitative methods for miRNAs with high specificity and sensitivity is a central challenge to make miRNA testing a routine part of medical care with respect to cancer. Herein, we propose a strategy for glucoamylase-encapsulated liposome-encoded magnetic beads initiated by padlock exponential rolling circle amplification (P-ERCA) for portable and accurate quantification of miRNA by using a glucometer readout.

DNA methyltransferase (MTase) plays a critical role in maintaining genome methylation patterns, which has a close relationship to cancer and bacterial diseases. This encouraged the need to develop highly sensitive, simple, and robust assays for DNA MTase detection and inhibitor screening. Herein, a simple, sensitive, and specific DNA MTase activity assay was developed based on magnetic beads-liposome hybrids combined with personal glucose meter (PGM) for quantitative detection of DNA MTase and inhibitor screening. First, a magnetic beads-liposome hybrid probe is designed by the hybridization of p 1 DNA-functionalized magnetic bead with p 2 DNA-functionalized glucoamylase-encapsulated liposome (GEL). It integrates target recognition, magnetic separation and signal amplification within one multifunctional design. Then, in the presence of Dam MTase, the hybrids probe was methylated, and cleaved by methylation-sensitive restriction endonuclease Dpn I, making liposome separated from magnetic bead by magnetic separation. Finally, the separated liposome was decomposed, liberating the encapsulated glucoamylase to catalyze the hydrolysis of the signal substrate amylose with multiple turnovers, producing a large amount of glucose for quantitative readout by the PGM. In the proposed assay, the magnetic beads-liposome hybrids offered excellent sensitivity due to primary amplification via releasing numerous glucoamylase from a liposome followed by a secondary enzymatic amplification. The use of portable quantitative device PGM bypasses the requirement of complicated instruments and sophisticated operations, making the method simple and feasible for on-site detection. Moreover, the proposed assay was successfully applied in complex biological matrix and screen suitable inhibitor drugs for DAM for disease(s) treatment. The results reveal that the approach provides a simple, sensitive, and robust platform for DNA MTases detection and screening potential drugs in medical research and early clinical diagnostics., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Developing a sensitive and selective sensing platform for the p53 gene and its mutation analysis is essential and may aid in early cancer screening and assessment of prognosis. Here, we developed a highly sensitive and selective p53 gene assay based on the coupling of a triple-helix magnetic probe (THMP) to a fluorescent liposome hybridization assembly, a process initiated by rolling circle amplification (RCA). In the presence of p53, the THMP unfolds and activates an enzymatic cleavage reaction, thus releasing the RCA primer and initiating the RCA product-assisted fluorescent liposome hybridization assembly. The resultant double-stranded DNA structures bind the intercalating SG dye from the fluorescent liposomes, thus dramatically enhancing the fluorescence signal. In the absence of p53, the THMP remains intact and blocks the trigger release and fluorescent liposome assembly, thus resulting in a low background signal. The THMPs were designed with integrated target recognition by Watson–Crick base-pairing, site-specific cleavage by an endonuclease and background signal elimination by magnetic isolation, thus avoiding the need to design multiple probes. Moreover, the use of fluorescent liposome assembly and magnetic isolation helps in avoiding sample matrix interference and nonspecific staining. Through cooperative amplification coupling with enzyme cleavage recycling, the RCA-assisted fluorescent liposome assembly and magnetic isolation improved the sensitivity, with a detection limit of 0.07 fM. The excellent capacity of the THMP to specifically detect the involved targets and the precise site-specific endonuclease cleavage ensured remarkable selectivity for p53 against single-base mismatches. This proposed approach worked well in biological samples, thus demonstrating great potential for biomedical and clinical diagnosis applications. [ABSTRACT FROM AUTHOR]

Here we developed a simple, sensitive and accurate PLD detection method based on a target-controlled gating liposome (TCGL) “off–on” cascade amplified strategy and personal glucose meters (PGMs). It showed excellent sensitivity with a detection limit of 0.005 U L−1 and well performed PLD activity analysis in breast cancer cells and inhibitor drug screening. [ABSTRACT FROM AUTHOR]