Bioelectrochemistry最新文献

MiR-1246 in breast cancer-derived exosomes was a promising biomarker for early diagnosis of breast cancer(BC). However, the low abundance, high homology and complex background interference make the accurate quantitative detection of miR-1246 facing great challenges. In this study, we developed an electrochemical biosensor based on the subtly combined of CRISPR/Cas12a, double-stranded specific nuclease(DSN) and magnetic nanoparticles(MNPs) for the detection of miR-1246 in BC-derived exosomes. Ascribed to the good synergistic effect of DSN, Cas12a and MNPs, the developed electrochemical biosensor exhibited excellent performance with the linear range from 500 aM to 5 pM, and the detection limit as low down to about 50 aM. The target-specific triggered enzyme-digest activity of DSN and Cas12a system, as well as the powerful separation ability of MNPs ensure the high specificity of developed electrochemical biosensor which can distinguish single base mismatches. In addition, the developed electrochemical biosensor has been successfully applied to detect miR-1246 in blood-derived exosomes and realize distinguishing the BC patients from the healthy individuals. It is expected that the well-designed biosensing platform will open up new avenues for clinical liquid biopsy and early screening of breast cancer, as well as provide deeper insights into clinical oncology treatment.

Intracellular reactive oxygen species (ROS) generation is widely suggested as a trigger for biological consequences of electric field exposures, such as those in electroporation applications. ROS are linked with membrane barrier function degradation, genetic damage, and complex events like immunological cell death. Dihydroethidium (DHE) is commonly used to monitor ROS in cells. DHE is linked to intracellular ROS by a primary oxidation product, Ethidium (Eth⁺), that shows increased fluorescence upon binding to polynucleotides. We observed changes in DHE-derived fluorescence in Chinese hamster ovary (CHO) cells post 300-ns electric pulse exposures, comparing them to tert-butyl-hydroperoxide (t-BHP) induced oxidative stress. Immediate intracellular fluorescence changes were noted in both cases, but with distinct localization patterns. After electrical stress, cytosolic DHE-derived fluorescence intensity decreases, and nucleolar intensity increases. Conversely, t-BHP exposure increases DHE-derived fluorescence uniformly across the cell. Surprisingly, fluorescence patterns after electrical stress in Eth⁺-loaded cells is identical to those in DHE-loaded cells, in kinetics and localization patterns. These findings indicate that DHE-derived fluorescence changes after pulsed electric field stress are not due to intracellular ROS generation leading to DHE oxidation, but rather indicate stress-induced intracellular microenvironment alterations affecting Eth⁺ fluorescence.

Overuse of enrofloxacin (ENR) has posed a potential threat to ecosystems and public health, so it is critical to sensitive and accurate determination of ENR residues. In this work, a novel ultra-sensitive and specific electrochemical aptasensor was fabricated based on the cobalt diselenide loaded gold and platinum nanoflowers (Au@Pt NFs/ CoSe₂) and Exonuclease III (Exo III)-assisted cycle amplification strategy for the detection of ENR. Au@Pt NFs/ CoSe₂ nanosheets as the substrate material, with large surface area, accelerate electron transfer and attach more DNA probes on the electrode substrate, have effectively enhanced the electrochemical performance of the electrode. With the existence of Enrofloxacin (ENR), the aptamer recognizes and binds to ENR, thus the signal probe cDNA was released and immobilized onto the electrode surface to hybridized with methylene blue (MB) labelled DNA (MB-DNA), thereby triggering the Exo III-assisted cycle for further signal amplification. As expected, the prepared aptasensor demonstrated excellent sensitivity and selectivity, with a wide linear range from 5.0 × 10⁻⁶ ng/mL to 1.0 × 10⁻² ng/mL for ENR, a low detection limit of 1.59 × 10⁻⁶ ng/mL. Consequently, this strategy provided a promising avenue for ultrasensitive and accurate detection of ENR in milk samples.

Carcinoembryonic antigen (CEA), a key colon biomarker, demands a precise detection method for cancer diagnosis and prognosis. This study introduces a novel electrochemical aptasensor using a triblock polyadenine probe for ultra-sensitive detection of CEA. The method leverages Exonuclease III (Exo III)-assisted target recycling and hybridization chain reaction. The triblock polyadenine probe self-assembles on the bare gold electrode through the strong affinity between adenine and gold electrode, blocking CEA diffusion and providing a large immobilization surface. CEA binding to hairpin probe 1 (HP1), followed by the hybridization between HP1 and hairpin probe 2 (HP2), triggers DNA cleavage by Exo III, amplifying the signal via a hybridization chain reaction and producing numerous dsDNA walkers that generates a dramatic electrochemical impedance signal. Under optimized conditions, the aptasensor achieved two ultra-low detection limits: 0.39 ag∙mL⁻¹ within the concentration range of 5 ag∙mL⁻¹ to 5 × 10⁶ ag∙mL⁻¹, and 1.5 ag∙mL⁻¹ within the concentration range of 5 × 10⁶ ag∙mL⁻¹ to 1 × 10¹⁰ ag∙mL⁻¹. Its performance in human serum samples meets the practical standards, offering a promising new tool for ultrasensitive tumor marker detection, potentially revolutionizing early cancer diagnosis.

In this study, we have designed an electrochemical biosensor based on topological material Bi₂Se₃ for the sensitive detection of SARS-CoV-2 in the COVID-19 pandemic. Flake-shaped Bi₂Se₃ was obtained directly from high-quality single crystals using mechanical exfoliation, and the single-stranded DNA was immobilized onto it. Under optimal conditions, the peak current of the differential pulse voltammetry method exhibited a linear relationship with the logarithm of the concentration of target-complementary-stranded DNA, ranging from 1.0 × 10^-15 to 1.0 × 10^-11 M, with a detection limit of 3.46 × 10^-16 M. The topological material Bi₂Se₃, with Dirac surface states, enhanced the signal-to-interference plus noise ratio of the electrochemical measurements, thereby improving the sensitivity of the sensor. Furthermore, the electrochemical sensor demonstrated excellent specificity in recognizing RNA. It can detect complementary RNA by amplifying and transcribing the initial DNA template, with an initial DNA template concentration ranging from 1.0 × 10^-18 to 1.0 × 10^-15 M. Furthermore, the sensor also effectively distinguished negative and positive results by detecting splitting-synthetic SARS-CoV-2 pseudovirus with a concentration of 1 copy/μL input. Our work underscores the immense potential of the electrochemical sensing platform based on the topological material Bi₂Se₃ in the detection of pathogens during the rapid spread of acute infectious diseases.

Lately, the bio electrochemical systems are emerging as an efficient wastewater treatment and energy conversion technology. However, their scaling-up is considerably restrained by slow-rate of cathodic oxygen reduction reaction (ORR) or otherwise by the high cost associated with the available efficient ORR catalysts. In this investigation, a cost-effective and eco-friendly approach for synthesizing Ni based ORR catalyst utilizing biosorption property of microalgae is accomplished. The synthesised Ni adsorbed algal biochar (NAB) served as an efficient cathode catalyst for enhancing ORR in a microbial carbon-capture cell (MCC). On increasing the initial concentration of Ni²⁺ in the aqueous medium from 100 mgL⁻¹ to 500 mgL⁻¹, the biosorption capacity was found to increase from 3 mgg⁻¹ to 32 mgg⁻¹ of algae cell. The MCC operated with NAB based cathode catalyst loading of 2 mgcm-² exhibited 3.5 times higher power density (4.69 Wm⁻³) as compared to the one with commercial activated carbon. A significant organic matter removal (82 %) in the anodic chamber with simultaneous algal biomass productivity in the cathodic chamber was attained by MCC with cathode loaded with 2 mgcm⁻² of NAB. Hence, this easily synthesised low-cost catalyst, out of waste stream, proved its ability to improve the performance of MCC.

Functional characterization of transporters is impeded by the high cost and technical challenges of current transporter assays. Thus, in this work, we developed a new characterization workflow that combines cell-free protein synthesis (CFPS) and solid supported membrane-based electrophysiology (SSME). For this, membrane protein synthesis was accomplished in a continuous exchange cell-free system (CECF) in the presence of nanodiscs. The resulting transporters expressed in nanodiscs were incorporated into proteoliposomes and assayed in the presence of different substrates using the surface electrogenic event reader. As a proof of concept, we validated this workflow to express and characterize five diverse transporters: the drug/H⁺-coupled antiporters EmrE and SugE, the lactose permease LacY, the Na⁺/H⁺ antiporter NhaA from Escherichia coli, and the mitochondrial carrier AAC2 from Saccharomyces cerevisiae. For all transporters kinetic parameters, such as K_M, I_MAX, and pH dependency, were evaluated. This robust and expedite workflow (e.g., can be executed within only five workdays) offers a convenient direct functional assessment of transporter protein activity and has the ability to facilitate applications of transporters in medical and biotechnological research.

Multiple sclerosis (MS) is a severe progressive autoimmune-inflammatory, demyelinating process in the central nervous system (CNS) with heterogeneous neurological symptoms appearing as a consequence of myelin break down. Myelin basic protein (MBP) makes up to 30 % of the CNS myelin [1] and it is known to be released into the cerebrospinal fluid (CSF) as a bioindicator of MS. Autoimmune encephalomyelitis (EAE) is a mice model of MS widely used for research and development of new treatments [2]. Herein, MBP specific aptamer developed for possible therapeutic purposes in mouse model [3] was applied as a bioreceptor for MBP recognition. A nanobiosensor for MBP detection and monitoring was developed by using graphene oxide (GO) nanoparticles integrated onto the screen-printed carbon electrodes (SPCE) and aptamer immobilized to create a bioactive layer on the sensor surface for MBP binding. The measurements were carried out using electrochemical impedance spectrometry (EIS). Validation studies were carried out in a biological matrix (artificial CSF) containing MBP, and MSA. The aptasensor had LOD in artificial CSF 0.01 ng/mL and showed its usability in the concentration range of 0.01 … 64 ng/mL.

The pathophysiological link between diabetes and heightened propensity for the development of coronary heart disease (CHD) is well-established. Prevailing evidence confirms that small increases in low concentrations of high-sensitivity C reactive protein (hs-CRP) in the human body can determine the tendency of developing CHD. Additionally, glycated hemoglobin (HbA1c) is a well-recognized biomarker to evaluate diabetes progression. Given the positive correlation between diabetes and CHD, this research presents a notably unprecedented label-free electrochemical approach for the dual detection of %HbA1c regarding Total Hb and hs-CRP, facilitating early CHD prediction and cost-effective point-of-care diagnostics. Furthermore, a novel redox probe O-(4-Nitrophenylphosphoryl)choline (C₁₁H₁₇N₂O₆P) was used for the electrochemical detection of CRP, a method not documented in scientific literature before. The calibration curves demonstrate a limit of detection (LOD) of 5 mg/mL in PBS (pH 8) and 6 mg/mL in simulated blood (SB) for a linear range of 0–30 mg/mL of HbA1c. Conjointly, a LOD of 0.007 mg/mL and 0.008 mg/mL for measurement in PBS (pH 7.4) and SB are reported for a linear range of 0–0.05 mg/mL of CRP. The electrochemical systems presented could accurately quantify HbA1c and CRP in mixed samples, demonstrating reasonable specificity and practical applicability for complex biological samples.

It is predicted that ultra-short electric field pulses (nanosecond) can selectively permeabilize intracellular structures (e.g., mitochondria) without significant effects on the outer cell plasma membrane. Such a phenomenon would have high applicability in cancer treatment and could be employed to modulate cell death type or immunogenic response. Therefore, in this study, we compare the effects of 100 µs x 8 pulses (ESOPE − European Standard Operating Procedures on Electrochemotherapy) and bursts of 100 ns pulses for modulation of the mitochondria membrane potential. We characterize the efficacies of various protocols to trigger permeabilization, depolarize mitochondria (evaluated 1 h  after treatment), the extent of ATP depletion and generation of reactive oxygen species (ROS). Finally, we employ the most prominent protocols in the context of Ca²⁺ electrochemotherapy in vitro. We provide experimental proof that 7.5–12.5 kV/cm x 100 ns pulses can be used to modulate mitochondrial potential, however, the permeabilization of the outer membrane is still a prerequisite for depolarization. Similar to 100 µs x 8 pulses, the higher the permeabilization rate, the higher the mitochondrial depolarization. Nevertheless, 100 ns pulses result in lesser ROS generation when compared to ESOPE, even when the energy input is several-fold higher than for the microsecond procedure. At the same time, it shows that even the short 100 ns pulses can be successfully used for Ca²⁺ electrochemotherapy, ensuring excellent cytotoxic efficacy.