Web hybrid chain reaction enhanced fluorescent magnetic bead array for digital nucleic acid detection.

The detection of biomarkers at low concentrations is important in clinical diagnostic analyses and has attracted continuous research. In this work, absolute quantification of hepatitis B virus (HBV) DNA was achieved using magnetic beads with isothermal, enzyme-free DNA nanostructure for fluorescence amplification. Firstly, the DNA-functionalized bead captured the target nucleic acid in the form of sandwich hybridization, and the individual target lighted up the entire bead by isothermal web hybridization chain reaction (wHCR). After the microarray scanning, the target nucleic acids can be digitally quantified based on the Poisson statistics. Therefore, the fluorescent bead assay enabled precise detection of HBV DNA down to 5 fM level without external calibration curves. Moreover, this method not only specifically distinguished single-base mismatched sequences, but also obtained the quantitative detection of HBV DNA in serum samples. Unlike routine digital detection usually combined with complex compartment partitioning operations, the amplification structure immobilized on beads can be conducted in microcentrifuge tubes with a volume of microliter scale. This work expands the application of magnetic beads in the digital quantitative detection via enzyme-free and isothermal method.

A Cascaded DNA Circuit in Bead Arrays for Quantitative Single-Cell MicroRNA Analysis.

Single-cell microRNA (miRNA) analysis helps people understand the causes of diseases and formulate new disease treatment strategies. However, miRNA from a single cell is usually very rare and requires signal amplification for accurate quantification. Here, to amplify the signal, we constructed the cascaded DNA circuits consisting of catalytic hairpin assembly and hybrid chain reaction into the bead array platform, on which the uniformly distributed beads were adopted for miRNA quantification. After exponential signal amplification, a consistent linear correlation between the percentage of fluorescent beads and the copy number of miRNA was detected. The proposed bead array can achieve ultrahigh sensitivity as low as 60 copies of miR-155 and high specificity for distinguishing single nucleotide differences. This method has been successfully applied to the quantitative detection of miRNA in a single cancer cell. The high sensitivity, programmability, and simple workflow of the bead array chip will give a huge advantage in basic and clinical research.

Investigation of the relationship between hot flashes, sweating and sleep quality in perimenopausal and postmenopausal women: the mediating effect of anxiety and depression.

BACKGROUND: To investigate the relationship between sweating from hot flashes, anxiety, depression, and sleep quality in peri- and postmenopausal women. And also the role of anxiety and depression in mediating sweating from hot flashes and sleep quality. METHODS: 467 women aged 40-60 years with menopausal problems were enrolled. The sleep quality; hot flashes; sweating; anxiety and depression symptoms were quantitatively evaluated by Pittsburgh Sleep Quality Scale (PSQI), Kupperman Menopause Index, Self-rating Anxiety Scale and Self-rating Depression Scale. Spearman correlation analysis and mediating effect model were used to analyze the relationship between the three. RESULTS: 262 patients' PSQI score were higher than 6 (58.2%). Total scores of sleep quality were positively correlated with hot flashes, sweating and anxiety and depression symptoms. Anxiety and depression played a mediating role between hot flashes, sweating and sleep quality where the mediating effect of anxiety symptoms accounted for 17.86% (P < 0.01) and depression symptoms accounted for 5.36% (P < 0.01). CONCLUSIONS: The hot flashes, sweating, anxiety and depression of peri/postmenopausal women are risk factors affecting sleep quality. By alleviating these risk factors, the sleep quality of peri- and postmenopausal women could be improved, which prevents the physical and mental diseases due to long-term severe insomnia.

A fluorescent microarray platform based on catalytic hairpin assembly for MicroRNAs detection.

The DNA microarray has distinctive advantages of high-throughput and less complicated operations, but tends to have a relatively low sensitivity. Catalytic hairpin assembly (CHA) is one of the most promising enzyme-free, isothermal DNA circuit for high efficient signal amplification. Here, a microarray-based catalytic hairpin assembly (mi-CHA) biosensing method has been developed to detect various miRNAs in a single test simultaneously. The target miRNA can trigger conformational transformations of hairpin-structured DNA probes on the chip surface and lead to the specific signal amplification. A significant advantage of this approach is that each duplex produced by the solid-phase CHA will be immobilized on the certain location of the chip and release fluorescent signal via the universal domain, eliminating the requirement of different fluorophores. This method has manifested a high detection sensitivity of human cancer-associated miRNAs (miR-21 and miR-155) down to 1.33 fM and promised a high specificity to distinguish single-base mismatches. Furthermore, the practicability of this method was demonstrated by analyzing target miRNAs in human serum and cancer cells. The experimental results suggest that the proposed method has high-throughput analytical potential and could be applied to many other clinical diagnosis.

Enzyme-free fluorescence microarray for determination of hepatitis B virus DNA based on silver nanoparticle aggregates-assisted signal amplification.

In this study, we designed a fluorescence enhancement strategy based on silver nanoparticle (AgNP) aggregates for the detection of hepatitis B virus DNA sequences. AgNPs were functioned with recognition probes (Cy3-probe) and hybrid probes (Oligomer-A and Oligomer-B). The presence of target DNA mediated the formation of sandwich complexes between the immobilized capture probes and the functionalized AgNPs, which was followed by hybridization-induced formation of AgNP aggregates. The fluorescent intensity could be extremely amplified by both the increasing number of fluorophores and metal enhanced fluorescence (MEF) effect. Under optimal conditions, this method achieved a detection limit of 50 fM which was 1560-fold lower than that of un-enhanced fluorescent assays. It was illustrated that the HBV DNA concentrations ranging from 100 fM to 10 nM had a good log-linear correlation with the corresponding fluorescent intensity (R = 0.991). Moreover, this method had high specificity both for distinguishing single-base mismatches and identifying target DNA under the interference of genomic DNA. This fluorescent microarray had high-throughput analytical potential and could apply to many other disease diagnoses.