Horseradish Peroxidase Research Articles

Immobilized Horseradish Peroxidase on Enriched Diazo-Activated Silica Gel Harnessed High Biocatalytic Performance at a Steady State in Organic Solvent.

Dimethyldichlorosilane (DMDCS), an efficient silane coupling reagent appearing between the -OH groups of silica gel (SG) and picric acid, instantaneously produces a derivative enriched with nitro groups. The nitro group acting as an end-cap terminates the reaction and subsequently was converted into diazo to couple tyrosine's phenol ring via its O-carbon, the inert center to immobilize horseradish peroxidase (HRP) in a multipoint mode. It maintains the status quo of the native enzyme's protein folding and the entire protein groups' chemistry. The molecular formula of the synthesized material was verified and appeared as {Si(OSi)4 (H2O)x}n{-O-Si(CH3)2-O-C6H2(N+≡N)3(HRP)}4·yH2O; the parameters were evaluated as x = 0.5, n = 1158, and y = 752. The immobilized biocatalyst's activity in organic solvents was 1.5 times better than that in an aqueous medium; it worked smoothly, wherein the activity in both solvents stabilized at six months and continued up to nine months at 63 ± 3% compared to the initial.

Biochemical properties of immobilized horseradish peroxidase on ceramic and its application in removal of azo dye.

In the current work, electrostatic interactions were used to immobilize the horseradish peroxidase (HRP) onto five types of ceramic materials (C) with different concentrations of oxidized metals (C1-C5). The highest immobilization efficiency (70 and 77%) was detected at 6mg C3 and 18 enzyme units. Scanning Electron Microscope (SEM), Energy Dispersive X-ray (EDX) and Fourier Transform Infrared (FTIR) analysis of C3-HRP confirmed the immobilization of the enzyme. After ten reuses, the reusability analysis showed that (66%) of the C3-HRP enzyme activity was retained. For C3-HRP, the optimum pH and temperature of the soluble enzyme were shifted from 7.0 and 30°C to 6.0 and 50°C. Up to 40°C and 50°C, respectively, the soluble HRP and C3-HRP remained steady. The kinetic analysis revealed that the Km and Vmax of soluble HRP and C3-HRP were, respectively, 5.5mM, 0.66 units, and 8mM, 0.52 units for hydrogen peroxide (H2O2) and 35.5mM, 3.4 units and 40mM, 1.1 units for guaiacol. Compared to soluble-HRP, the C3-HRP exhibited a greater oxidizing affinity toward several phenolic compounds (Guaiacol, o-dianisidine, o-phenylenediamine, pyrogallol, p-aminoantipyrine). In comparison with soluble-HRP, the C3-HRP showed increased stress tolerance with Triton X-100, urea, metals, isopropanol, and dimethyl sulfoxide. The C3-HRP removed methyl orange more effectively compared to soluble-HRP.

Rapid Microfluidic Biosensor for Point-of-Care Determination of Rheumatoid Arthritis via Anti-Cyclic Citrullinated Peptide Antibody Detection

Rheumatoid arthritis (RA) is a chronic autoimmune disorder that causes extensive damage to multiple organs and tissues and has no known cure. This study introduces a microfluidic detection platform that combines a microfluidic reaction chip with a micro-spectrometer to accurately detect the anti-cyclic citrullinated peptide antibody (anti-CCP Ab) biomarker, commonly associated with arthritis. The surface of the microfluidic reaction chip is functionalized using streptavidin to enable the subsequent immobilization of biotinylated-labeled cyclic citrullinated peptide (biotin–CCP) molecules through a streptavidin–biotin reaction. The modified chip is then exposed to anti-CCP Ab, second antibody conjugated with horseradish peroxidase (HRP) (2nd Ab-HRP), 3,3′,5,5′-tetramethylbenzidine (TMB), and a stop solution. Finally, the concentration of the anti-CCP Ab biomarker is determined by analyzing the optical density (OD) of the colorimetric reaction product at 450 nm using a micro-spectrometer. The detection platform demonstrated a strong correlation (R2 = 0.9966) between OD and anti-CCP Ab concentration. This was based on seven control samples with anti-CCP Ab concentrations ranging from 0.625 to 100 ng/mL. Moreover, for 30 artificial serum samples with unknown anti-CCP Ab concentrations, the biosensor achieves a correlation coefficient of (R2 = 0.9650). The proposed microfluidic detection platform offers a fast and effective method for accurately identifying and quantifying the anti-CCP Ab biomarker. Thus, it offers a valuable tool for the early diagnosis and monitoring of RA and its progression in point-of-care settings.

Spatiotemporal Proximity-Enhanced Biocatalytic Cascades Within Metal-Organic Frameworks for Wearable and Theranostic Applications.

Enzymatic catalysis, particularly multi-enzyme cascade catalytic, is often limited by the spatial and temporal separation of enzymes and their signal substrates. Herein, a facile method for producing a spatiotemporal proximity-enhanced biocatalytic cascade system is introduced by encasing enzymes within metal-organic frameworks (MOFs) that are modulated with sulfonic acid-functionalized signal substrates. The modulated behavior relies on the sulfonic acid groups coordinated with Zn2+. As a proof of concept, by utilizing 2,2'-Azinobis (3-ethylbenzothiazoline-6-sulfonic acid ammonium salt) (ABTS), a widely-used signal substrate for horseradish peroxidase, two-enzyme/substrate, and three-enzyme/substrate MOFs, which demonstrated a 7.4- and 10.2-fold increase in biocatalytic efficiency over free systems are successfully synthesized. Incorporating the synthesized MOFs into homemade wearable patches and in vivo settings, noninvasive sweat glucose colorimetric detection and photoacoustic imaging-guided photothermal tumor therapy are enabled, respectively. This advancement stems from the newly established coordinative bonds between Zn2+ centers and substrates' sulfonic acid groups, which negates the need for additional signal substrates, thereby not only enhancing but also streamlining bioapplication processes.

Mathematical Modeling and Optimization of Enzyme‐Based Amperometric Screen‐Printed Biosensors Using Horseradish Peroxidase as a Model

AbstractIn this study, a mathematical model was formulated to characterize the functioning of third‐generation enzyme‐based amperometric biosensors. This study utilized a horseradish peroxidase (HRP)‐based, screen‐printed biosensor. The model parameters were determined using the Environment for Modeling Simulation and Optimization (EMSO), validated with experimental data, and exhibited high determination coefficients, indicating the accuracy of the models in capturing biosensor behavior. Simulations based on this model emphasize diffusion as a limiting factor. Further sensitivity analysis was conducted using simulated response surfaces and biosensor performance optimization, which confirmed that higher concentrations of HRP in the thinnest polymeric film resulted in enhanced biosensor sensitivity.

Plasma Deposition of Parylene-like Films with Chemical Functional Groups for Immunoassays.

Parylene-like films obtained via the plasma decomposition of parylene precursors with functional groups (amino and formyl) are proposed as an alternative to those obtained via the thermal method. To analyze the chemical functional groups after plasma deposition, a surface analysis of the parylene films using the two different deposition methods was performed via Fourier transform infrared (FT-IR) and X-ray photoelectron spectroscopy (XPS). The FT-IR analysis revealed that the featured peaks of the chemical functional groups were maintained in the parylene-like films obtained via the plasma deposition method. The XPS analysis revealed that the featured chemical functional groups of parylene-AM and parylene-H were maintained after plasma deposition. The surface energy of the parylene films was estimated by using contact angle measurements. The plasma-deposited parylene films were then employed for protein immobilization via the functional groups using horseradish peroxidase (44 kDa) and green fluorescent protein (25 kDa) as model proteins. The parylene-AM and parylene-H films obtained via plasma deposition exhibited higher immobilization efficiencies than did the same parylene films obtained via thermal deposition. Finally, a competitive immunoassay was obtained by immobilizing the Fv-antibodies on plasma-deposited parylene-AM and parylene-H films via covalent bonding. Using heat-deactivated SARS-CoV-2 as a real sample, the limit of detection at the feasible level for the medical diagnosis of COVID-19 was achieved using a competitive immunoassay based on immobilized Fv-antibodies on plasma-deposited parylene films.

MgAl-Layered Double Hydroxide-Coated Bio-Silica as an Adsorbent for Anionic Pollutants Removal: A Case Study of the Implementation of Sustainable Technologies.

The adsorption efficiency of Cr(VI) and anionic textile dyes onto MgAl-layered double hydroxides (LDHs) and MgAl-LDH coated on bio-silica (b-SiO2) nanoparticles (MgAl-LDH@SiO2) derived from waste rice husks was studied in this work. The material was characterized using field-emission scanning electron microscopy (FE-SEM/EDS), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and X-ray photoelectron spectroscopic (XPS) techniques. The adsorption capacities of MgAl-LDH@SiO2 were increased by 12.2%, 11.7%, 10.6%, and 10.0% in the processes of Cr(VI), Acid Blue 225 (AB-225), Acid Violet 109 (AV-109), and Acid Green 40 (AG-40) dye removal versus MgAl-LDH. The obtained results indicated the contribution of b-SiO2 to the development of active surface functionalities of MgAl-LDH. A kinetic study indicated lower intraparticle diffusional transport resistance. Physisorption is the dominant mechanism for dye removal, while surface complexation dominates in the processes of Cr(VI) removal. The disposal of effluent water after five adsorption/desorption cycles was attained using enzymatic decolorization, photocatalytic degradation of the dyes, and chromate reduction, satisfying the prescribed national legislation. Under optimal conditions and using immobilized horseradish peroxidase (HRP), efficient decolorization of effluent solutions containing AB-225 and AV-109 dyes was achieved. Exhausted MgAl-LDH@SiO2 was processed by dissolution/precipitation of Mg and Al hydroxides, while residual silica was used as a reinforcing filler in polyester composites. The fire-proofing properties of composites with Mg and Al hydroxides were also improved, which provides a closed loop with zero waste generation. The development of wastewater treatment technologies and the production of potentially marketable composites led to the successful achievement of both low environmental impacts and circular economy implementation.

A novel colorimetric assay for the detection of urinary N1, N12-diacetylspermine, a known biomarker for colorectal cancer

Urinary N1, N12-diacetylspermine (DAS) is a known biomarker for colorectal cancer (CRC). However, DAS levels in both healthy and CRC patients’ urine samples are extremely low and often challenging to quantify. Complex and expensive methods do exist to detect DAS in urine, but simpler, less expensive methods to detect DAS are needed, especially in low resource settings. Here we describe a highly efficient, fast, precise, and inexpensive colorimetric assay to detect low levels of DAS in human urine samples. We used recombinant diacetylspermine oxidase (rDAS Ox), expressed and extracted from E. coli, to oxidize DAS, producing three products including hydrogen peroxide (H2O2). The level of DAS present, which correlates with H2O2 levels, was measured using horseradish peroxidase (HRP), which together with H2O2, oxidized Amplex™ Red to produce the pink-colored resorufin. The concentration of resorufin is directly proportional to H2O2 (and DAS) levels. As urine contains metabolites which interfere with these oxidation reactions, we developed a simple two column-based protocol using ion exchange resins to remove these compounds and concentrate the DAS. With this novel cleaning and concentrating method, DAS was concentrated 15 times (confirmed by nuclear magnetic resonance (NMR) spectroscopy) and <1 μM DAS could be detected. Correlation graphs of urine samples spiked with known DAS concentrations versus assay-determined DAS concentrations had high coefficients of determination (R2) for 0-10 μM DAS (0.94) and for 0-1 μM DAS (0.91), clearly demonstrating the excellent performance of the two-column protocol with the rDAS Ox reaction mixture. To the best of our knowledge, this is first reported colorimetric enzymatic assay that quantitates DAS in urine.

Chemical Synthesis and Enzymatic Modification of Mangostins: A Comprehensive Review on Structural Modifications for Drug Discover.

Mangosteens, a prominent component of Garcinia mangostana, have been ex-tensively studied for their biological activities and structural modifications. Chemical methods, including cyclization reactions under acidic conditions, have yielded many de-rivatives, which often exhibit enhanced pharmacological properties compared to itself. Enzymatic biotransformation, such as glycosylation and oxidation mediated by fungal species and enzymes like horseradish peroxidase, have provided regioselective pathways to functionalized mangostin derivatives. These studies highlight the versatility of mangos-tin as a scaffold for designing compounds with tailored biological functions. Overall, mangosteen represents a promising platform for developing compounds with enhanced pharmacological activities, paving the way for innovative approaches in biomedicine and pharmaceutical sciences. This review provides a comprehensive examination of the chem-istry of mangosteens, detailing their total synthesis and the derivatives obtained through both chemical and enzymatic methodologies.

A dual-aptamer sandwich assay by detection of c-reactive protein in synovial fluid on an integrated microfluidic system for diagnosis of periprosthetic joint infection

Diagnosing periprosthetic joint infection (PJI) in equivocal cases is always challenging. Synovial fluid (SF) C-reactive protein (CRP) levels have been reported as a surrogate to improve the diagnosis accuracy. This study reported a new integrated microfluidic system (IMS) designed to detect SF-CRP at a dynamic range of 1-50mg/L. The novel, magnetic bead-based, dual-aptamer assay with two CRP-specific aptamers, St1 and St2, enabled rapid and accurate CRP detection. This represents the first attempt to automatically perform the CRP dual-aptamer assay on a microfluidic chip quantifying CRP in SF with optical signals. CRP concentrations in fourteen clinical samples were accurately quantified by using the dual-aptamer assay on the developed IMS in an automatic format, suggesting the assay’s potential as a reliable diagnostic tool for SF-CRP detection and may serve as an armamentarium for PJI diagnosis. Notably, the assay was deemed superior to traditional enzyme-linked immunosorbent assay (ELISA) because it consumes a smaller amount of sample (only 40μL), requires fewer reagents (only 50μL of secondary detection aptamer and horseradish peroxidase), and provides quicker results (~2hours).

Construction of hierarchical porous MIL-100(Fe) for horseradish peroxidase immobilization and its degradation performance of bisphenol A

Construction of hierarchical porous MIL-100(Fe) for horseradish peroxidase immobilization and its degradation performance of bisphenol A

Real-time simultaneous visualization of lactate and proton dynamics using a 6-μm-pitch CMOS multichemical image sensor

Multi-analyte detection and imaging of extracellular chemical signaling molecules are crucial for understanding brain function and molecular pathology. In this work, we present a 6-μm-pitch, CMOS-based multichemical image sensor that enables the simultaneous visualization and spatiotemporal multimodal analysis of the lactate and proton (H+) dynamics without any labeling. Using semiconductor lithography, gold electrode patterns functioning as lactate-sensing regions were formed on a potentiometric sensor array. Lactate is detected potentiometrically because of redox reactions using lactate oxidase and horseradish peroxidase. The resulting multichemical image sensor exhibited a pH sensitivity of 65 mV and a superior detection limit of 1 μM for lactate with a reasonable selectivity. Furthermore, diffusion images of lactate and H+ distributions were obtained concurrently, allowing for simultaneous real-time imaging of the two chemicals with subcellular resolution. We believe that our novel imaging device can be successfully applied to extracellular microenvironments in tissue or cell samples as an effective bioimaging tool.

Construction of Liposome‐Based Extracellular Artificial Organelles on Individual Living Cells

AbstractIntegration of living cells with extrinsic functional entities gives rise to bioaugmented nanobiohybrids, which hold tremendous potential across diverse fields such as cell therapy, biocatalysis, and cell robotics. This study presents a biocompatible method for incorporating multilayered functional liposomes onto the cell surface, creating extracellular artificial organelles or exorganelles. The introduction of various extrinsic functionalities to cells is achieved without comprising their viabilities. The integration of extrinsic enzymatic reactions is exemplified through the cascade reaction involving glucose oxidase and horseradish peroxidase. Furthermore, our protocol offers the design flexibility to customize liposome compositions, thereby providing effective cell modification. The versatility of the liposome‐based exorganelle approach establishes an advanced chemical tool, empowering cells with novel functionalities that surpass or are complementary to their innate capabilities.

Quantitative Intracellular Delivery of Anticancer Nanodrugs Via an Immunoassay Employing Pt-SiO2 Janus-Peroxidase Nanozyme.

The accurate and efficient quantification of nanodrug dosage is crucial for early anticancer therapy. The enzyme-linked immunosorbent assay (ELISA) has emerged as a robust tool for detecting anticancer nanodrug dosage; however, the development of sensing elements to quantify anticancer nanodrugs still poses a challenge. To overcome this problem, we utilize polysuccinimide-loaded curcumin (CUR @PSIOAm) as a model to employ an ELISA based on peroxidase nanozyme Pt-SiO2 Janus nanoparticles (Pt-SiO2 JNPs) for the indirect quantitative analysis of intracellular anticancer nanodrug dosage. This novel approach employs an immunoassay to indirectly quantify the dosage of anticancer nanodrugs while preserving its structural integrity. The silica components of Pt-SiO2 JNPs adsorb intermediates, while the Pt NP components exhibit high catalytic activity. Pt-SiO2 JNPs are functionalized with anti-PSIOAm antibody (Pt-SiO2 JNPs-Ab) to serve as an immunosensor capable of specific recognition of CUR @PSIOAm. Additionally, we employed cytotoxicity assays and confocal imaging techniques to demonstrate the excellent biocompatibility of CUR @PSIOAm, as well as its specific uptake by cancer cells. According to the experimental results, the limit of detection (LOD) for the immunoassay of Pt-SiO2 JNPs as a marker for detecting CUR @PSIOAm is approximately 4.5-fold lower than that of horseradish peroxidase. Therefore, by optimizing the conditions, we established a direct competitive ELISA using Pt-SiO2 JNPs as colorimetric indicators for the quantitative detection of intracellular CUR @PSIOAm. The LOD for this ELISA was determined to be 0.01 ng/mL, while the loaded CUR amount calculated from the drug loading capacity was found to be 0.22 pg/mL. Furthermore, the recoveries obtained from this established ELISA ranged between 94.0 and 108%, demonstrating excellent accuracy. Consequently, the peroxidase mimic Pt-SiO2 JNPs-based ELISA exhibits significant potential for precise quantification of intracellular anticancer nanodrug dosages.

Fusarium graminearum Ste2 and Ste3 Receptors Undergo Peroxidase-Induced Heterodimerization when Expressed Heterologously in Saccharomyces cerevisiae.

Fusarium graminearum FgSte2 and FgSte3 are G-protein coupled receptors (GPCRs) shown to play roles in hyphal chemotropism and fungal plant pathogenesis in response to activity arising from host-secreted peroxidases. Here, we follow up on the observation that chemotropism is dependent on both FgSte2 and FgSte3 being present; testing the possibility that this might be due to formation of an FgSte2-FgSte3 heterodimer. Bioluminescence resonance energy transfer (BRET) analyses were conducted in Saccharomyces cerevisiae, where the addition of horse radish peroxidase (HRP) was found to increase the transfer of energy from the inducibly-expressed FgSte3-Nano luciferase donor, to the constitutively-expressed FgSte2-yellow fluorescent protein (YFP) acceptor, compared to controls. A partial response was also detected when an HRP-derived ligand-containing extract was enriched from F. graminearum spores and applied instead of HRP. In contrast, substitution with pheromones or an unrelated bovine GPCR, rhodopsin-YFP used as acceptor, eliminated all BRET responses. Interaction results were validated by affinity pulldown and receptor expression was validated by confocal immunofluorescence microscopy. Taken together these findings demonstrate the formation of HRP and HRP-derived ligand stimulated heterodimers between FgSte2 and FgSte3. Outcomes are discussed from the context of the roles of ligands and reactive oxygen species in GPCR dimerization.

Unlocking of Hidden Mesopores for Enzyme Encapsulation by Dynamic Linkers in Stable Metal-Organic Frameworks.

Mesoporous metal-organic frameworks (MOFs) are promising supports for the immobilization of enzymes, yet their applications are often limited by small pore apertures that constrain the size of encapsulated enzymes to below 5 nm. In this study, we introduced labile linkers (4,4',4''-(2,4,6-boroxintriyl)-tribenzoate, TBTB) with dynamic boroxine bonds into mesoporous PCN-333, resulting in PCN-333-TBTB with enhanced enzyme loading and protection capabilities. The selective breaking of B-O bonds creates defects in PCN-333, which effectively expands both window and cavity sizes, thereby unlocking hidden mesopores for enzyme encapsulation. Consequently, this strategy not only increases the adsorption kinetics of small enzymes (<5 nm) such as cytochrome c (Cyt C) and horseradish peroxidase (HRP), but also enables the immobilization of various large-sized enzymes (>5 nm), such as glycoenzymes. The glycoenzymes@PCN-333-TBTB platform was successfully applied to synthesize thirteen complex oligosaccharides and polysaccharides, demonstrating high activity and enhanced enzyme stability. The dynamic linker-mediated enzyme encapsulation strategy enables the immobilization of enzymes exceeding the inherent pore size of MOFs, thus broadening the scope of enzymatic catalytic reactions achievable with MOF materials.

Unlocking of Hidden Mesopores for Enzyme Encapsulation by Dynamic Linkers in Stable Metal‐Organic Frameworks

AbstractMesoporous metal‐organic frameworks (MOFs) are promising supports for the immobilization of enzymes, yet their applications are often limited by small pore apertures that constrain the size of encapsulated enzymes to below 5 nm. In this study, we introduced labile linkers (4,4′,4′′‐(2,4,6‐boroxintriyl)‐tribenzoate, TBTB) with dynamic boroxine bonds into mesoporous PCN‐333, resulting in PCN‐333‐TBTB with enhanced enzyme loading and protection capabilities. The selective breaking of B−O bonds creates defects in PCN‐333, which effectively expands both window and cavity sizes, thereby unlocking hidden mesopores for enzyme encapsulation. Consequently, this strategy not only increases the adsorption kinetics of small enzymes (&lt;5 nm) such as cytochrome c (Cyt C) and horseradish peroxidase (HRP), but also enables the immobilization of various large‐sized enzymes (&gt;5 nm), such as glycoenzymes. The glycoenzymes@PCN‐333‐TBTB platform was successfully applied to synthesize thirteen complex oligosaccharides and polysaccharides, demonstrating high activity and enhanced enzyme stability. The dynamic linker‐mediated enzyme encapsulation strategy enables the immobilization of enzymes exceeding the inherent pore size of MOFs, thus broadening the scope of enzymatic catalytic reactions achievable with MOF materials.

Enhanced Peroxidase-like Activity of Ruthenium-Modified Single-Atom-Thick A Layers in MAX Phases for Biomedical Applications.

Nanozymes have demonstrated significant potential as promising alternatives to natural enzymes in biomedical applications. However, their lower catalytic activity compared to that of natural enzymes has limited their practical utility. Addressing this challenge necessitates the development of innovative enzymatic systems capable of achieving specific activity levels of natural enzymes. In this study, we focus on enhancing the catalytic performance of nanozymes by introducing Ru atoms into the single-atom-thick A layer of the V2SnC MAX phase, resulting in the formation of V2(Sn0.8Ru0.2)C with Ru single-atom sites. The V2(Sn0.8Ru0.2)C MAX phase demonstrated an exceptional peroxidase-like specific activity of up to 1792.6 U mg-1, surpassing the specific activity of a previously reported horseradish peroxidase (HRP). Through X-ray photoelectron spectroscopy (XPS) and density functional theory (DFT) investigations, it has been revealed that both the V2C atom layers and single-atom-thick Sn readily accept a negative charge from Ru, leading to a reduction of the energy barrier for H2O2 adsorption. This discovery has enabled the successful application of V2(Sn0.8Ru0.2)C in the development of a lateral flow immunoassay for heart failure biomarkers, achieving a detection sensitivity of 4 pg mL-1. Additionally, V2(Sn0.8Ru0.2)C demonstrated exceptional broad-spectrum antibacterial efficacy. This study lays the groundwork for the precise design of MAX phase-based nanozymes with high specific activity, offering a viable alternative to natural enzymes for various applications.

Distance-based analytical device integrated with carbon nanomaterials for sarcosine quantification in human samples.

Astraightforward distance-based paper analytical device (dPAD) was developedfor monitoring sarcosine levels in human samples for the rapid diagnosis and prognosis of prostate cancer and related symptoms. This assay eliminates the need for the expensive horseradish peroxidase (HRP) enzyme by utilizing carbon nanodots (CDs) as a peroxidase-like nanozyme. The proposed dPAD sensor consists of a sample zone pre-deposited with sarcosine oxidase (SOx) and CDs, and a detection zone containing 3,3',5,5'-tetramethylbenzidine (TMB). When a solution containing sarcosine is added to the sample zone, hydroxyl radicals (•OH) are produced through SOx oxidation and subsequent peroxidase catalysis by the CDs. The formed •OH radicals immediately flow to the detection zone via capillary force, where they oxidize TMB, resulting in a visible colour change from colourless to blue. Sarcosine quantification is effortlessly accomplished by measuring the distance of the blue colour in the detection zone. The developed dPAD offers a linear working range between 12.5 and 35.0nmol L-1 (R2 = 0.9959) and a detection limit (LOD) of 10.0nmol L-1. This covers the clinical range for urinary sarcosine determination, thereby suggesting no additional sample preparation or dilution is needed. The sensor shows high precision with the highest relative standard deviation (RSD) of 4.58% and demonstrates excellent selectivity with no observed interferences. Furthermore, recovery studies in human control urine samples ranged from 98.67 to 101.50%, with the highest RSD of 2.03%. Correspondingly, our dPAD method showed a great match with the performance of a commercial ELISA method for detecting sarcosine in human control serum. The sensor is more cost-effective, user-friendly, and accessible than previous methods. Overall, the proposed method represents a promising analytical tool for sarcosine quantification. The concept is also applicable for broader analytical applications in detecting other biomolecules.

Aptamer-Based Electrochemical Biosensing Platform for Analysis of Cardiac Biomarkers.

Monitoring biomarkers secreted by cardiomyocytes is critical to evaluate anticancer drug-induced myocardial injury (MI). Cardiac troponin I (cTnI) is considered the gold standard biomarker for MI. Herein, an electrochemical aptasensor is engineered for cTnI detection based on lanthanide europium metal-organic frameworks (Eu-MOFs) and a hybridization chain reaction-directed DNAzyme strategy. Three types of Eu-MOF morphologies were easily synthesized by changing the solvent, and the Eu-MOF modulated by mixing the solvent of dimethylformamide and H2O (D-Eu-MOF) exhibited the best performance compared to other morphologies of the Eu-MOFs. Multifunctional nanoprobes were constructed from D-Eu-MOF@Pt loaded with natural horseradish peroxidase and combined with an aptamer-initiated nuclear acid hybridization chain reaction to form G-quadruplex/hemin DNAzymes for signal amplification. A novel capture probe is constructed on the basis of DNA nanotetrahedrons modified on screen-printed gold electrodes to enhance the capture of the target and multifunctional nanoprobes for signal amplification. It exhibits a detection limit of 0.17 pg mL-1 and a linear range from 0.5 pg mL-1 to 15 ng mL-1. The practicality of the platform is evaluated by measuring cTnI in real samples and secreted by cardiomyocytes after drug treatment, which provides great potential in drug-induced MI evaluation for clinical application.