A dual-core 3D DNA nanomachine based on DNAzyme positive feedback loop for highly sensitive MicroRNA imaging in living cells.

A double 3D DNA walker nanomachine by DNAzyme self-driven positive feedback loop amplification for the detection of miRNA was constructed. This method uses two gold nanoparticles as the reaction core, and because of the spatial confinement effect the local concentration of the reactants increase the collision efficiency was greatly improved. Meanwhile, the introduction of positive feedback loop promotes the conversion efficiency. In presence of miRNA-21, a large amount of DNAzyme was released and hydrolyze the reporter probe, resulting the recovery of fluorescence signal. The linear range for miRNA-21 is 0.5-60 pmol/L, and the detection limit is 0.41 pmol/L (S/N = 3). This nanomachine has been successfully used for accurate detection of miRNA-21 expression levels in cell lysates. At the same time, it can enter cells for intracellular miRNA-21 fluorescence imaging, distinguishing tumor cells from normal cells. This combination of in vitro detection and imaging analysis of living cells can achieve the goal of jointly detecting cancer markers through multiple pathways, providing new ideas for early diagnosis and screening of diseases.

Bimetallic FeOx-TiO2@Carbon hybrid structure materials with notable peroxidase enzyme mimics applied to one-step colorimetric detection of glucose.

FeOx-TiO2@Carbon hybrid structure materials (FeOx-TiO2@CHs) with high peroxidase (POD)-like activity have been prepared by one-pot hydrothermal method. Based on the excellent POD activity of FeOx-TiO2@CHs, one pot colorimetric detection for glucose was constructed by using TMB as substrate with the synergistic reaction of glucose oxidase; the linear range and the limit of detection (LOD) are 25 ~ 1000 and 1.77 µM, respectively. Using this method, the glucose in serum real samples was detected with satisfactory results, and the results are consistent with that of the glucometer method in the hospital. The recovery in diabetic and artificial urine samples was 95.71 ~ 104.67% and 99.01 ~ 103.16%, respectively. The mechanism of the catalytic colorimetric reaction was also investigated by multiple measurements, and the results indicated that superoxide anions (O2•-) between FeOx-TiO2@CHs and substrate play a main role, but a small quantity of hydroxyl radical •OH and singlet oxygen 1O2 is also generated simultaneously. The one-pot reaction method is simple and fast; the detection process only requires a simple mixing, which is suitable for application in special environment.

Simple enzyme-free detection of uric acid by an in situ fluorescence and colorimetric method based on Co-PBA with high oxidase activity.

In this work, we prepared a simple and low-cost cobalt-doped Prussian blue analog (Co-PBA), which can directly oxidize 10-acetyl-3,7-dihydroxyphenoxazine and 3,3',5,5'-tetramethylbenzidine (TMB) to produce resorufin (ox-AR) with high fluorescent quantum yield and ox-TMB with blue color, respectively, without the need for unstable H2O2. Using the Michaelis-Menten curve and Lineweaver-Burk equation, the Michaelis-Menten constant of Co-PBA and the substrate TMB was found to be 0.033 mM, which was much lower than horseradish peroxidase and other reported nanozymes, showing satisfactory substrate affinity. Uric acid (UA) can cause erosion of the Co-PBA structure, and it significantly reduces the catalytic activity of Co-PBA, resulting in the decrease of the fluorescence emission signal of ox-AR and the absorption signal of ox-TMB. Based on this, a simple, sensitive, and fast fluorescence/colorimetric dual-mode uric acid detection platform was established. The detection range for UA by fluorescence method is 0.625-40 µM, and the detection limit (LOD, S/N = 3) is as low as 0.389 µM. The detection system was applied to serum samples with good recovery and can be used for field detection of UA in biological samples under different environments to meet different needs.

Optimizing the method for removing MSNs templates using an ionic liquid ([C4mim]Cl).

The key step in preparing mesoporous silica is to remove the organic template agent, and the most common method used to achieve this goal is high-temperature calcination. However, this method has many disadvantages, one of which is that it reduces the silanol density on the surface of mesoporous silica, which affects its subsequent modification. Ionic liquids (ILs) are often used as extractants. In this work, the 1-butyl-3-methylimidazolium chloride ([C4mim]Cl) IL is considered, and the effects of its concentration, reaction temperature, and reaction time as well as HCl concentration on the extraction rate and silanol density were investigated using an IL extraction template agent (cetyl trimethyl ammonium bromide (CTAB)). The results show that an IL concentration of 10%, a reaction temperature of 120 °C, a reaction time of 12 h, and an HCl concentration of 1% are the best reaction parameters; with these parameters, the extraction rate and the silanol density were found to be 93.19% and 2.23%, respectively. The silanol density of mesoporous silica treated by calcination is only 0.81%. A higher silanol density provides more reaction sites, so that the modified mesoporous silica treated with the IL can be loaded with more Zn ions.

The Characteristics and Influencing Factors of Nonsuicidal Self-Injury of Adolescents With Depressive Disorder in China: A Meta-analysis.

ABSTRACT: This study is a systematic review of characteristics and influencing factors of nonsuicidal self-injury behavior among adolescents with depressive disorder in China. PubMed, CNKI, WanFang Database, and VIP were searched for studies. Meta-analysis was performed using RevMan 5.1 software. Nineteen studies involving 707 subjects were included in the meta-analysis. Age, gender, only child or not, and residence were included in the analysis, of which age ( I2 = 0%, p = 0.42) and residence ( I2 = 0%, p = 0.84) were analyzed by fixed-effects model; gender ( I2 = 75%, p = 0.003) and only child or not ( I2 = 50%, p = 0.140) were analyzed by random-effects model. The evidence shows that, according to the common self-injury mode and location, the nonsuicidal self-injury behavior of young people with depressive disorder aged 15-18 years is paid attention to and guided, so as to achieve early detection and early diagnosis and treatment, and reduce the occurrence of serious harm.

A novel nanoparticle surface-constrained CRISPR-Cas12a 3D DNA walker-like nanomachines for sensitive and stable miRNAs detection.

The CRISPR-Cas system has broad prospects as a new type of nucleic acid signal amplification technology based on the trans-cleavage activity of Cas12a to single-stranded DNA, but the trans-cleavage reaction efficiency is relatively low in solution. In order to overcome this negative factor, a new 3D DNA nanomachine whose CRISPR-Cas12a is limited to the surface of nanoparticles is used for sensitive and stable detection of miRNA. By loading Cas12a activator onto spherical nucleic acid (SNA), the CRISPR-Cas12a activator system on the surface of Au nanoparticles (AuNPs) acts as a walker to carry out continuous recognition-walking-cutting reaction on the surface of AuNPs, which enhances the trans-cleavage activity of Cas12a to SNAs. Benefiting from the confinement effect of spherical nucleic acids surface, a 3D DNA nanomachine has been developed for the detection of miRNA-21, which has achieved high sensitivity and accuracy, and the detection limit is able to reach 8.0 pM. This new 3D DNA walker-like nanomachine provided another insight for future bioanalysis and early clinical diagnoses of disease and liquid biopsy.

A novel label-free universal biosensing platform based on CRISPR/Cas12a for biomarker detection.

The development of a biosensing platform with high sensitivity, high specificity, and low cost for the detection of biomarkers, especially one that is programmable and universal, is critical for disease surveillance and diagnosis, yet it remains a difficulty. Herein, we combined the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system with a fluorescent label-free biosensor platform for sensitive and specific detection of disease-related protein, small molecule and nucleic acid. In this strategy, we designed an exonuclease III-mediated target cycle and released a universal trigger chain to stimulate the enzyme activity of CRISPR/Cas12a for additional signal amplification. The hydrolysis of ssDNA-templated silver nanoclusters (ssDNA-Ag NCs) as the reporter probe resulted in a significant decrease of fluorescence intensity. This biosensing platform can be flexibly used to the sensitive and specific determination of protein, small molecule, or microRNA in biological samples by simply transforming the target recognized sequences in the DNA hairpin. In this work, a new label-free sensing system used the fluorescent ssDNA-Ag NCs as the signal output does not need to be marked in advance and has no background signal. In addition, the method has the advantages of low cost, simple operation and high speed, and provides an innovative idea for the development of a powerful clinical diagnosis tool.

A DNAzyme-driven random biped DNA walking nanomachine for sensitive detection of uracil-DNA glycosylase activity.

Highly specific and ultrasensitive detection of uracil-DNA glycosylase (UDG) activity is of great significance for maintaining genomic integrity and medical research of related diseases. Here, we constructed a random DNA walking nanomachine based on a DNAzyme for UDG activity detection on the AuNP (Au nanoparticle) surface. When UDG is present, the U bases in the Y structure are removed, resulting in AP sites, which will be cleaved by Endo-IV to generate a 3' concave end for Exo-III, causing the locking strand of the DNAzyme to be completely hydrolyzed by the Exo-III and release the walking strand to randomly pair with the substrate strand on the AuNP surface; then, the walking strand exerts its cleavage activity with the assistance of Mg2+ to cleave the substrate strand and keep the fluorophore 6-carboxyfluorescein (FAM) away from the surface of the AuNP, which restores the fluorescence signal of this system. In this way, sensitive detection of UDG can be realized, and the detection limit is as low as 3.69 × 10-6 U mL-1. In addition, we found that this method is highly specific to UDG and can be used to detect UDG specifically in complex samples, which has certain application prospects in biomedical research and clinical diagnosis related to UDG.

Ratiometric fluorescent 3D DNA walker and catalyzed hairpin assembly for determination of microRNA.

Using 6-carboxyfluorescein (FAM) and tetramethyl rhodamine (TAMRA) as fluorescent signals a ratiometric fluorescent three-dimensional (3D) DNA walker based on a catalytic hairpin assembly (CHA) reaction for microRNA-122 detection was constructed. This method uses CHA reaction triggered indirectly by the target to mediate the 3D DNA walker operation to amplify the signal. The dual emission ratio fluorescent signal with a single excitation wavelength was used as the signal output. This strategy combines DNA walker with CHA reaction and proportional fluorescence signal output methods, which can effectively reduce the background fluorescence signal and the risk of generating false-positive signals. Thus, the impact of environmental factors on the experiment is reduced, thereby obtaining reliable and stable experimental results. It uses the fluorescence excitation wavelength of 488 nm and the maximum fluorescence emission wavelength of 520 nm and 580 nm, respectively. It has a good linear response at a microRNA concentration range of 156.0 pM ~ 7.00 nM and a detection limit of 42.94 pM. This strategy has been successfully applied to detect microRNAs in spiked serum samples. Graphical abstract Schematic representation of three-dimensional (3D) DNA walker constructed using catalytic hairpin self-assembly reaction (CHA)-assisted amplification and ratiometric fluorescence signal output for the detection of miRNA-122 closely related to hepatitis.

Ultrasensitive photoelectrochemical biosensor for MiRNA-21 assay based on target-catalyzed hairpin assembly coupled with distance-controllable multiple signal amplification.

Here, with the target-catalyzed hairpin assembly generated dsDNA (HP1-HP2) to synchronously control the departure of quencher ferrocene and approach of sensitizer methylene blue, a distance-controllable multiple signal amplification based photoelectrochemical biosensor was proposed for MiRNA-21 assay.

A dual-label time-resolved fluorescence immunoassay for the simultaneous determination of carcinoembryonic antigen and squamous cell carcinoma antigen.

Among women worldwide, cervical cancer is the second-most common cancer, and cervical smears and DNA detection have low sensitivity or are too expensive. The concentrations of carcinoembryonic antigen (CEA) and squamous cell carcinoma (SCC) in the serum were detected using a sandwich immunoassay. The CEA and SCC in the serum were captured by anti-CEA and anti-SCC antibodies. After combining other anti-CEA- and anti-SCC-labeled antibodies with europium (III) (Eu3+ ) and samarium (III) (Sm3+ ) chelates, CEA and SCC were detected with time-resolved fluorometry (TRF). The linear correlation coefficients (R2 ) of the CEA and SCC standard curves were 0.9997 and 0.9997, respectively. The minimum detection level for CEA was 1.15 ng/mL (the linear dynamic range was 3.24-543.67 ng/mL), and the average recovery was 100.83%. The sensitivity for SCC detection was 0.54 ng/mL (the linear dynamic range was 2.47-96.58 ng/mL), and the average recovery was 101.02%. High R2 between the results of commercial assays and this method were obtained (R2  = 0.9983 for CEA, R2   = 0.9878 for SCC). These findings indicated that the dual-label TRFIA invented in this study has high sensitivity, accuracy, and specificity in clinical analysis, which indicates that this method could be used for the early diagnosis and follow-up surveillance of cervical cancer.

CE with chemiluminescence detection for the determination of thyroxine in human serum.

A sensitive and rapid approach to perform thyroxine (T4) assay by CE with chemiluminescence (CL) detection was developed. The sensitive detection was based on the enhancement effect of T4 on the CL reaction between luminol and potassium permanganate (KMnO4 ) in alkaline solution. A laboratory-built reaction flow cell and a photon counter were deployed for the CL detection. Experimental conditions for CL detection were studied in detail to achieve maximum assay sensitivity. Optimal conditions were found to be 5.0 × 10(-4) M luminol added to the CE running buffer and 9.2 × 10(-5) M KMnO4 in 0.072 M NaOH solution introduced postcolumn. In the optimized experimental conditions, the linear range for T4 detection was 6.0 × 10(-8) -6.0 × 10(-6) M, with the detection limit of 2.0 × 10(-8) M (S/N = 3). Six human serum samples from healthy subjects, hyperthyroid patients and hypothyroid patients were analyzed by the presented method. The serum level of T4 in healthy subjects was found be 9.0 × 10(-8) M, whereas the T4 level was found to be 15.6 × 10(-8) M in hyperthyroid patients and 1.3 × 10(-8) M in hypothyroid patients. The results suggested a potential application of the proposed assay in rapid primary diagnosis of diseases such as hyperthyroid and hypothyroid.

Use of capillary electrophoresis with chemiluminescence detection for sensitive determination of homocysteine.

A sensitive capillary electrophoresis (CE) method with chemiluminescence (CL) detection was developed for the determination of homocysteine (HCys) in human plasma. In this work, N-(4-aminobutyl)-N-ethylisoluminol was used as tagging reagent to label the analyte for achieving high assay sensitivity. N-(4-Aminobutyl)-N-ethylisoluminol-tagged HCys after CE separation reacted with hydrogen peroxide in the presence of horseradish peroxidase, producing CL emission. Experimental conditions for labeling analyte, CE separation, and CL detection were studied. The CL intensity was proportional to the concentration of HCys in the range of 2.5×10(-8) to 5.0×10(-6) M. Detection limit (S/N=3) was 7.6×10(-9) M. Human plasma samples from healthy donors were analyzed by the presented method. HCys levels were found to be in the range of 9.50-15.3 µM.

Preparation and properties of poly(vinyl alcohol)-stabilized liposomes.

The purpose of this work is to evaluate the improvement in physical stability of poly(vinyl alcohol) (PVA) modified liposomes. Liposomes composed of soya phosphatidylcholile (SPC) and cholesterol (1:1 molar ratio) were prepared by reverse phase evaporation method. Two types of interaction between liposome and PVA were investigated: PVA addition into lipid bilayer during liposome preparation and coating of already formed liposomes with PVA. The microparticles system was morphologically characterized by transmission electron microscopy (TEM) and particles analysis. Changes in particles size and zeta potential confirmed the existence of a thick polymer layer on the surface of liposomes. The amount of PVA adsorbing to liposomes and the encapsulation efficiency increased with increasing polymer concentration. The physical stability was evaluated by measuring the release rate of contents at 20 and 37 degrees C, the PVA modified liposomes were more stable than the conventional liposomes. Comparing with PVA-coated liposomes, the liposomes with PVA addition to the bilayer were more stable, and had higher entrapment efficiency.