Portable Detection of Lysine Acetyltransferase Activity in Lung Cancer Cells Based on a Miniature Electrochemical Sensor.

The detection of lysine acetyltransferases is crucial for diagnosing and treating lung cancer, highlighting the necessity for highly efficient detection methods. We developed a portable, highly accurate, and sensitive technique using fast-scan cyclic voltammetry (FSCV) to determine the activity of the lysine acetyltransferase TIP60, employing a novel miniature electrochemical sensor. This approach involves a compact electrochemical cell, merely 3 mm in diameter, that holds solutions up to 50 µL, equipped with a conductive indium tin oxide working electrode. Uniquely, this system operates on a two-electrode model compatible with the FSCV, obviating the traditional requirement for a reference electrode. The system detects TIP60 activity through the continuous generation of CoA molecules that engage in reactions with Cu(II), thereby significantly improving the accuracy of the acetylation analysis. Remarkably, the detection limit achieved for TIP60 is notably low at 3.3 pg/mL (S/N = 3). The results show that the reversible dynamic acetylation can be effectively regulated by inhibitor incubation and glucose stimulation. This cutting-edge strategy enhances the analysis of a broad spectrum of biomarkers by modifying the responsive unit, and its miniaturization and portability significantly amplify its applicability in biomedical research, promising it to be a versatile tool for early diagnostic and therapeutic interventions in lung cancer.

Dual-network hydrogels based on dynamic imine and borate ester bonds with antibacterial and self-healing properties.

Polymeric hydrogel materials with multiple functions are in great demand in practical biomedical scenarios. In this work, a self-healing hydrogel with both antimicrobial properties was prepared using a strategy that combines dynamic imine and borate ester bonds. In this hydrogel, polyvinyl alcohol (PVA) is used as the base network, and borax solution as the cross-linking agent, and borate ester bonds can be formed between these two. Dialdehyde carboxymethyl cellulose (DCMC) was selected to cross-link with the amino groups in carboxymethyl chitosan (CMCS) and polyethyleneimine (PEI) to form dynamic imine bonds. The PVA/PEI/DCMC/CMCS hydrogels prepared by double chemical cross-linking have good mechanical properties (maximum tensile strength up to 289 KPa and strain at the break up to 1025%). Due to the uniqueness of the two chemical bonds, the hydrogel material is self-healing at room temperature without additional stimulation. In addition, the inherent antibacterial properties of the raw materials in this hydrogel confer antibacterial properties, with a kill rate of up to 99% against E. coli and S. aureus. The multifunctional hydrogels developed in this study provide more ideas and references for the future application of hydrogel materials in practical scenarios.

Electrochemical sensor for single-cell determination of bacteria based on target-triggered click chemistry and fast scan voltammetry.

Herein, an electrochemical sensor for single-cell determination of bacteria was developed based on target-triggered click chemistry and fast scan voltammetry (FSV). In it, bacteria not only are the detection target, but also can use their own metabolism to achieve first-level signal amplification. More electrochemical labels were immobilized on functionalized 2D nanomaterials to achieve second-level signal amplification. At 400 V/s, FSV can achieve third-level signal amplification. The linear range and limit of quantification (LOQ) are 1 âˆ¼ 108 CFU/mL and 1 CFU/mL, respectively. When the reaction time of E. coli-instructed Cu2+ reduction is extended to 120 min, PCR-free single-cell determination of E. coli was achieved by electrochemical method first time. The feasibility of the sensor was verified by analysis of E. coli in seawater and milk samples with recoveries ranging from 94% to 110%. This detection principle is widely applicable, providing a new path for the establishment of single-cell detection strategy for bacteria.

The effects of glyphosate exposure on gene transcription and immune function of the silkworm, Bombyx mori.

Glyphosate is a widely used herbicide and crop desiccant. However, whether its extensive use has any effect on the species diversity of nontarget organisms is still unclear. In this study, we used the silkworm, Bombyx mori, as the research subject, and performed RNA sequencing to analyze the transcriptional profile of silkworm midgut after exposure to glyphosate at 2975.20 mg/L (a concentration commonly used at mulberry fields). A total of 125 significantly differentially expressed genes (DEGs) were detected in the midgut of glyphosate-exposed silkworm (q < 0.05), of which 53 were upregulated and 72 were downregulated. Gene ontology enrichment analysis showed that the DEGs were mainly enriched in biological process, cellular component, and molecular function. Kyoto encyclopedia of genes and genomes analysis showed that the differential genes were mainly related to oxidative stress, nutrient metabolism, and immune defense pathways, including oxidative stress-related Cat and Jafrac1, nutrient metabolism-related Fatp and Scpx, and immune-related CYP6AN2, UGT40B4, CTL11, serpin-2, and so forth. Experimental verification showed that glyphosate exposure led to a 4.35-fold increase in the mortality of silkworm after Beauveria bassiana infection, which might be caused by the decreased PO (phenoloxidase) activity and impaired immunity. These results provide evidence for the potential effects of residue glyphosate on the physiological functions of silkworm, and also provide a reference for the biosafety evaluation of glyphosate.

Checkerboard arranged G4 nanostructure-supported electrochemical platform and its application to unique bio-enzymes examination.

In this study, a checkerboard arranged G4 nanostructure-supported electrochemical platform is developed well for the application to unique bio-enzymes examination. Herein, we focus on the two bio-enzymes involving histone acetyltransferase (HAT) and terminal deoxynucleotidyl transferase (TdT); the former leads to the acetyl transfer of acetyl coenzyme A to the lysine residue of the substrate peptide and the latter achieves the polymeric extension of DNA without template under a unique pool of dATP and dGTP (4: 6). A complex of antibody and short DNA is introduced onto the electrode surface based on the affinity interaction between acetyl in acetylated peptide and its antibody. and used for initiating reaction. Then, TdT-yielded rich-G sequence is formed to act as the backbone of checkerboard, and subsequently free G-DNAs can be stacked continually on the backbone under Mg2+. An excellent electrocatalytic signal to H2O2 emerges noticeably, which is related with the activity of HAT p300 and TdT with a low detection limit of 2.3 pM and 0.38 mU/mL, respectively. Finally, this strategy is also used successfully for the inhibitor screening and complex sample analysis, which has important implications for the advancement of HAT- and TdT-related electrochemical bioassay and drug discovery.

Development of ZnO nanoflowers-assisted DNAzyme-based electrochemical platform for invertase and glucose oxidase-dominated biosensing.

The investigation on invertase (INV) and glucose oxidase (GOx)-dominated biological process offers a new opportunity for the development of clinical diagnosis and prognostic treatment. Herein, a ZnO nanoflowers (ZnONFs)-assisted DNAzyme-based electrochemical platform for INV- and GOx-dominated biosensing is proposed by the change of pH in microenvironment. In this strategy, INV can usually catalyze the dissolution of sucrose to generate glucose, and glucose is then consumed by GOx to produce H2O2 and gluconic acid, in which ZnONFs can be effectively etched into free Zn2+ ions. Subsequently, the released Zn2+ ions have a shearing action for Zn2+-specific DNAzyme, thus triggering hybridization chain reaction along with the imbedding of methylene blue. The excellent electrochemical signals illustrate the method can be employed well for testing sucrose, INV and GOx with a low detection limit (0.019 µM, 0.047 mU/mL and 0.012 mU/mL, respectively). Finally, a series of basic and advanced logic gates (YES, AND, INHIBIT, and AND-AND-INHIBIT) in the biological process are constructed with different logic inputs, providing a valuable platform for the establishment of advanced molecular devices for bioanalysis and clinical diagnostics.

Tunneling or Hopping? A Direct Electrochemical Observation of Electron Transfer in DNA.

We developed an axis-mode donor-DNA-acceptor electrochemical system to distinguish whether electron transfer in DNA occurs by tunneling or hopping. In the axis-mode, rigid stem-loop DNA was designed with the redox probe Ag+ embedded at the axis of the strand through a C-Ag+-C mismatch, which was immobilized onto the electrode surface in a saturated manner. Thus, the rotation, swing, and bending of the DNA strand were restricted and then the number of Ag+, the distance L between Ag+ and the electrode, and the chemical environment could be precisely controlled. In addition, fast scan cyclic voltammetry was applied to realize the in situ redox reaction of Ag+, without diffusion away from the electrode and the ensuing deconstruction of the stem-loop DNA. In this case, as a direct indicator of rate, the peak Faradaic current ip was extracted and used to fit the tunneling mechanism i ∝ exp (-ßL) and the hopping mechanism i ∝ L-η. The value of ß was determined to be 0.100 Å-1, which is consistent with the range of 0.1∼1.5 Å-1 reported previously, while η was determined to be 0.677, which is completely beyond the correct range of 1 ≤ η ≤ 2, demonstrating that electron transfer in DNA occurs by tunneling instead of hopping or that tunneling dominates. Additionally, current additivity and the irrelevance of the base sequence illustrate this point again. Thus, the possibility of independent parallel tunneling currents in DNA strands is revealed, which is helpful for recognizing the feasibility of DNA-based wires and devices.

Protonation-induced DNA conformational-change dominated electrochemical platform for glucose oxidase and urease analysis.

Cytosine and protonated cytosine base pairs (C·CH+)-supported i-motif conformation has been widely employed in some interdisciplinary fields such as biology, medicine and chemistry. In this work, we report a new electrochemical biosensing method for the detection of glucose oxidase (GOx) and urease based on pH-induced DNA conformational-change. The constructed platform mainly includes TdT-mediated catalytic synthesis, GOx- or urease-catalyzed biological reaction and pH-induced DNA conformational-change. In the beginning, a kind of C-rich DNA is produced by TdT catalysis, and multiple C·CH+-supported i-motif structures appear under acidic condition. Then, the oxidation of glucose catalyzed by GOx or the hydrolyzation of urea aroused by urease can result in a generation of acidic or alkaline environment owing to the generated gluconic acid or ammonia. Herein, protonation and deprotonation interaction in TdT-yielded C-rich DNA can lead to different electrochemical impedance spectroscopy (EIS) toward Fe(CN)63-/4-. Based on it, the EIS response changes proportionally toward GOx concentrations from 0.01 to 20 U/L or urease concentrations from 0.01 to 50 U/L, and the detection limit of GOx or urease is 0.0061 U/L or 0.0028 U/L (S/N = 3), respectively. Beyond this, we also construct a series of molecular logic gates (YES, AND, NOT, and NAND) with good performance by altering inputs under long C-rich DNA substrate. These excellent properties indicate that the unique sensing platform is potential to monitor GOx or urease in practical biosystems and clinical medical examinations.

[miR-152 inhibits the epithelial-mesenchymal transition and renin-angiotensin system of human hepatocellular carcinoma cells by down-regulating AGTR1].

Objective To investigate the effects of microRNA-152 (miR-152) targeting at angiotensin II type 1 receptor (AGTR1) on the epithelial mesenchymal transition (EMT) and renin-angiotensin system (RAS) of HCCLM3 human hepatocellular carcinoma cells. Methods The cultured HCCLM3 cells were divided into untransfected group (untreated), negative control group (transfection negative control sequence) and miR-152 group (transfected miR-152 mimic). The expressions of miR-152, angiotensin converting enzyme (ACE), angiotensin II (AngII) and angiotensin II type 1 receptor (AGTR1) mRNAs were detected by real-time fluorescence quantitative PCR. Cell invasion and migration were detected by TranswellTM assay. The expression of vimentin, N-cadherin, E-cadherin and AGTR1 were tested by western blot. The targeting relationship between miR-152 and AGTR1 were examined by double luciferase reporter assay. Results Compared with the untransfected group or the negative control group, the expression levels of miR-152 and E-cadherin protein in the miR-152 group significantly increased, while the expression levels of ACE, AngII, AGTR1 mRNA, the number of invaded cells, the number of migrating cells, and the protein expression levels of vimentin, N-cadherin, and AGTR1 decreased significantly. The results of double luciferase reporter gene assay confirmed that miR-152 can target binding with AGTR1. Conclusion miR-152 may inhibit EMT and RAS of HCCLM3 cells by targeting down-regulation of AGTR1 expression.

Polyvinyl alcohol/carboxymethyl chitosan hydrogel loaded with silver nanoparticles exhibited antibacterial and self-healing properties.

Hydrogel materials are gradually increasing research in biological aspects due to their unique properties. In order to prepare hydrogels with the potential to be used in clinical wound therapy, the authors prepared a bifunctional hydrogel with antibacterial and self-healing properties. The hydrogel was composed of borax cross-linked polyvinyl alcohol (PVA) and carboxymethyl chitosan (CMCS), which realizes self-healing between polymers through hydrogen bonds and borate ester bonds. The double cross-linking of hydrogen bonds and borate ester bonds also endows the hydrogel with better mechanical properties (toughness and tensile stress can reach 22.30 MJ/m3 and 70.35 KPa, respectively). On this basis, adding highly stable silver nanoparticles (AgNPs) to the hydrogel can effectively inhibit the growth of E. coli and S. aureus. This idea provides the possibility for the application of hydrogels in the process of biological wound healing.

Electrochemical aptasensor for Staphylococcus aureus by stepwise signal amplification.

An electrochemical aptasensor for ultrasensitive detection of Staphylococcus aureus (SA) has been developed based on stepwise signal amplification. In the sample processing stage, the specific recognition between SA and aptamer triggers the enzyme-assisted cyclic cleavage to produce a large amount of target DNA (tDNA), realizing the first-level signal amplification. In the sensor assembly stage, tDNA induces a catalytic hairpin assembly (CHA) cycle to capture much more hairpin DNA H2 labeled by the electrochemical tag ferrocene, bringing the second-level signal amplification. In the signal detection stage, ferrocene is quasi-adsorbed on the electrode surface, and a high redox peak current linearly increasing with the scan rate up to 1000 V/s has been obtained by fast scan cyclic voltammetry (FSCV), achieving the third-level signal amplification. Under the optimized experimental conditions, the linear range and detection limit are 1 ~ 108 CFU/mL and 0.3 CFU/mL, respectively. The sensor has good reproducibility, stability, and sensitivity, affording practical sample detection. This detection principle is widely applicable to other pathogens, and provides a new path for the establishment of highly sensitive detection strategies.

Supramolecular Host-Guest Interaction-Driven Electrochemical Recognition for Pyrophosphate and Alkaline Phosphatase Analysis.

We report an electrochemical biosensor based on the supramolecular host-guest recognition between cucurbit[7]uril (CB[7]) and L-phenylalanine-Cu(II) complex for pyrophosphate (PPi) and alkaline phosphatase (ALP) analysis. First, L-Phe-Cu(II) complex is simply synthesized by the complexation of Cu(II) (metal node) with L-Phe (bioorganic ligand), which can be immobilized onto CB[7] modified electrode via host-guest interaction of CB[7] and L-Phe. In this process, the signal of the complex-triggered electro-catalytic reduction of H2 O2 can be captured. Next, due to the strong chelation between PPi and Cu(II), a biosensing system of the model "PPi and Cu(II) premixing, then adding L-Phe" was designed and the platform was applied to PPi analysis by hampering the formation of L-Phe-Cu(II) complex. Along with ALP introduction, PPi can be hydrolyzed to orthophosphate (Pi), where abundant Cu(II) ions are released to form L-Phe-Cu(II) complex, which gives rise to the catalytic reaction of complex to H2 O2 reduction. The quantitative analysis of H2 O2 , PPi and ALP activity was achieved successfully and the detection of limits are 0.067 µM, 0.42 µM and 0.09 mU/mL (S/N=3), respectively. With its high sensitivity and selectivity, cost-effectiveness, and simplicity, our analytical system has great potential to for use in diagnosis and treatment of ALP-related diseases.

Pt nanoclusters embedded Fe-based metal-organic framework as a dual-functional electrocatalyst for hydrogen evolution and alcohols oxidation.

Design and construction of high-efficiency and durable dual-functional electrocatalyst for clean energy electrocatalytic reaction is urgently desirable for mitigating the energy shortage and environmental deterioration issues. Herein, we prepared Pt nanoclusters with exposed (111) face plane embedded Fe-based metal-organic frameworks (Fe-MOF, MIL-100(Fe)) catalyst for electrocatalytic hydrogen evolution reaction (HER) and ethylene glycol oxidation reaction (EGOR). It is noted that the available oxygen sites on the surface of MIL-100(Fe) would form Pt-O interaction with Pt nanoclusters to acquire strong interfacial interaction, which endows Pt/MIL-100(Fe) electrocatalyst effective electron transfer, increasing catalytic active sites, accelerating proton-electron coupling, and improving conductivity. Benefitting from the desirable metal-supports interaction and derive merits for catalysis, the high electrocatalytic activity and durability for HER and EGOR were achieved as expected. Impressively, superior HER performance with higher current density, lower overpotential (46/29 mV in acidic/alkaline electrolyte) and smaller Tafel slope (19.7/37.8 mV dec-1 in acidic/alkaline electrolyte) were acquired compared to commercial Pt/C. Moreover, Pt/MIL-100(Fe) electrode exhibits a rather high mass activity of 11826 mA mg-1Pt and long-term stability for EGOR. The present investigation demonstrates the promise of active metal/MOF combination for the interfacial strategy and rational design of dual-functional electrocatalyst, which has potential applications for future electrocatalysis field.

Cascade i-motifs-dependent reversibleelectrochemical impedance strategy-oriented pH and terminal deoxynucleotidyl transferase biosensing.

In this study, we develop a novel and reversibleelectrochemical impedance strategy for pH and terminal deoxynucleotide transferase (TdT) analysis based on the TdT-assisted generation of long enough cytosine (C)-rich DNAs. The formation of this special DNA is rationally designed on 5'-thiol DNA modified Au electrode surface, and TdT can catalyze the extension of this 3'-OH end to form a long C-rich DNA in the presence of deoxycytidine triphosphate (dCTP). Here, we discover a reversible process, in which the TdT-generated C-rich DNA maintains an irregular single chain state under neutral conditions and some stable DNA i-motifs (cascade i-motifs) are formed due to the partial protonation of C under acidic conditions. More importantly, the electrochemical impedance spectroscopy (EIS) response varies with the configuration change of the TdT-mediated C-rich DNA under different pH conditions. In view of this, a unique EIS switch ("on-off-on") is constructed faithfully with the configuration change, thus achieving pH analysis well. Additionally, the TdT activity can be also detected well by recording the EIS response, because it can catalyze the DNA tailing process up to hundreds of cytosines; on the contrary, if its inhibitor exists, TdT-based extension and formation of cascade i-motifs will not occur. Using this strategy, the detection of limit for TdT is 0.79 × 10-5 U/mL (pH 7.0) and 0.25 × 10-5 U/mL (pH 5.8) (S/N = 3), respectively. All the above features make our biosensor a promising assay for in situ monitoring of pH and TdT in complex clinical diagnosis.

A ratiometric fluorescence sensor based on carbon quantum dots realized the quantitative and visual detection of Hg2.

In this paper, based on the fluorescence of carbon quantum dots (CQDs) quenched by mercury ions (Hg2+ ) and the nonresponse of Hg2+ to rhodamine B fluorescence, a dual emission ratio fluorescence sensor was constructed to realize the quantitative detection of Hg2+ . Under excitation at 365 nm, the fluorescence spectrum showed double emission peaks at 437 nm and 590 nm, corresponding to the fluorescence emissions of CQDs and rhodamine B, respectively. This method quantitatively detected Hg2+ based on the linear relationship between the ratio of the intensities of the two emission peaks F437 /F590 and the concentration of Hg2+ . The detection range was 10-70 nM, and the limit of detection (S/N = 3) was 3.3 nM. In addition, this method could also realize the qualitative and semiquantitative detection of Hg2+ according to the fluorescence colour change of the probe under ultraviolet light. After various evaluations, the method could be successfully applied to the quantitative and visual detection of Hg2+ in tap water, and demonstrated excellent selectivity, anti-interference performance, and repeatability of the method.

DNA walker-mediated biosensor for target-triggered triple-mode detection of Vibrio parahaemolyticus.

Herein, we have constructed a target-triggered and DNA walker-mediated biosensor with triple signal (BTS) outputs mode for sensitive and reliable detection of pathogenic bacteria. Vibrio parahaemolyticus (VP) being the detection target model, the aptamer conformational changes induced by VP have been designed to activate the DNA walk on the modifiable and conductive surface of Fe3O4 nanoparticles to generate triple signal outputs, including electrochemiluminescence (ECL), fast scan cyclic voltammetry (FSCV) and fluorescent pixel counting (FLPC). Limits of quantification (LOQ) of VP were as low as 1 CFU⋅mL-1 by ECL with a linear range of 1-106 CFU⋅mL-1, 1 CFU⋅mL-1 by FSCV with a linear range of 1-106 CFU⋅mL-1, and 10 CFU⋅mL-1 by FLPC with a linear range of 10-107 CFU⋅mL-1 respectively, all squared correlation coefficients R2 being > 0.99. In addition, optical and electrochemical results, signal-on and signal-off results, electrode phase and solution phase results could be mutually verified by integrating of multiple detection techniques in one biosensor, greatly improving the accuracy and reliability. Therefore, the designed BTS has provided a powerful strategy for pathogenic bacteria detection considering its high detection sensitivity and accuracy, exhibiting great potential in food safety, water quality and biological contamination.

Faraday cage-type aptasensor for dual-mode detection of Vibrio parahaemolyticus.

A Faraday cage-type aptasensor has been developed for dual-mode detection of a common bacterial pathogen Vibrio parahaemolyticus (VP) by electrochemiluminescence (ECL) and differential pulse voltammetry (DPV), using a multi-functionalized material Pb2+-Ru-MOF@Apt2 as signal unit. The recognition aptamer Apt2 recognizes VP; specifically, ruthenium-based metal organic framework (Ru-MOF) and lead ions (Pb2+) embedded produce an ECL signal at a working potential from 0 to 1.5 V and DPV signal from - 0.3 to - 0.8 V vs. Ag/AgCl. Since Ru-MOF is a two-dimensional conductive material signal unit overlapped onto the electrode surface to form a Faraday cage-type aptasensor. Thus, electrons could be easily exchanged between electrode and signal tags without being hindered by micron-size VP, resulting in a high detection sensitivity with a detection limit of 1.7 CFU mL-1. In addition, dual-mode detection was achieved, improving the accuracy and reliability of rapid field detection. Stability and selectivity were also satisfactory. The tests of real samples indicate that this Faraday cage-type aptasensor is suited for rapid detection of VP and analog pathogens and shows great potential in food safety. Graphical abstract.

Reprogrammable fluorescence logic sensing for biomolecules via RNA-like coenzyme A-based coordination polymer.

In this study, coenzyme A (CoA)-based coordination polymers (CPs) have been generated in situ by exploiting the reaction of thiols with metal ion (Au(III) or Ag(I)), which are dependent on both thiol-metal and aurophilic metal∙metal interaction. It is interesting to note that CPs-related biosensing capabilities toward some biomolecules including ascorbic acid (AA), cysteine (Cys) and glutathione (GSH) are also investigated via SYBR Green II (SGII)-derived fluorescence switchable mechanisms. The synthesized CPs display especial RNA-like structure and are capable of initiating the fluorescence of SGII. Conversely, AA, Cys or GSH can give rise to the structural destruction of RNA-like CPs, thus inhibiting the fluorescence signal, and quantitative detection of these biomolecules are achieved favorably with a detection limit of 7.2, 0.55 and 0.48 nM, respectively. Meanwhile, the fascinating fluorescence on-off property and simple synthetic process are employed to build a series of basic logic gates (YES, NOT, OR, AND, INHIBIT and NOR) and multiple configurable logic gates (OR-AND and OR-OR-INHIBIT) along with different logic inputs. In view of these, developing CoA-based CPs as a new material to execute logic operations provides a valuable platform to establish the next generation of advanced molecular devices for clinic diagnostic and biomedical research.