Graphene quantum dots on TiO2 nanotubes as a light-assisted peroxidase nanozyme.

Hybrid nanozyme graphene quantum dots (GQDs) deposited TiO2 nanotubes (NTs) on titanium foil (Ti/TiO2 NTs-GQDs) were manufactured by bestowing the hybrid with the advantageous porous morphology, surface valence states, high surface area, and copious active sites. The peroxidase-like activity was investigated through the catalytic oxidation of chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H2O2, which can be visualized by the eyes. TiO2 NTs and GQDs comprising oxygen-containing functional groups can oxidize TMB in the presence of H2O2 by mimicking peroxidase enzymes. The peroxidase-mimicking activity of hybrid nanozyme was significantly escalated by introducing light illumination due to the photosensitive features of the hybrid material. The peroxidase-like activity of Ti/TiO2 NTs-GQDs enabled H2O2 determination over the linear range of 7 to 250 µM, with a LOD of 2.1 µM. The satisfying peroxidase activity is possibly due to the unimpeded access of H2O2 to the catalyst's active sites. The porous morphology provides the easy channeling of reactants and products. The periodic structure of the material also gave rise to acceptable reproducibility. Without material functionalization, the Ti/TiO2 NTs-GQDs can be a promising substitute for peroxidases for H2O2 detection.

Realizing Ultrasensitive and Accurate Point-of-Care Profiling for ATP with a Triple-Mode Strategy Based on the ATP-Induced Reassembly of a Copper Coordination Polymer Nanoflower.

New strategies for accurate and reliable detection of adenosine triphosphate (ATP) with portable devices are significant for biochemical analysis, while most recently reported approaches cannot satisfy the detection accuracy and independent of large instruments simultaneously, which are unsuitable for fast, simple, and on-site ATP monitoring. Herein, a unique, convenient, and label-free point-of-care sensing strategy based on novel copper coordination polymer nanoflowers (CuCPNFs) was fabricated for multimode (UV-vis, photothermal, and RGB values) onsite ATP determination with high selectivity, sensitivity, and accuracy. The resulting CuCPNFs with a 3D hierarchical structure exhibit the ATP-triggered decomposition behavior because the competitive coordination between ATP and the copper ions of CuCPNFs can result in the formation of ATP-Cu, which reveals preeminent peroxidase mimics activity and can accelerate the oxidation of 3, 3', 5, 5'-tetramethylbenzidine (TMB) to form oxTMB. During this process, the detection system displayed not only color changes but also a strong NIR laser-driven photothermal effect. Thus, the photothermal and color signal variations are easily monitored by a portable thermometer and a smartphone. This multimode point-of-care platform can meet the requirements of onsite, without bulky equipment, accuracy, and reliability all at once, greatly enhancing its application in practice and paving a new way in ATP analysis.

Flexible microfluidic colorimetric detection chip integrated with ABTS·+ and Co@MnO2 nanozyme catalyzed TMB reaction systems for bio-enzyme free detection of sweat uric acid.

BACKGROUND: The development of wearable detection devices that can achieve noninvasive, on-site and real-time monitoring of sweat metabolites is of great demand and practical significance for point-of-care testing and healthcare monitoring. Monitoring uric acid (UA) content in sweat provides a simple and promising way to reduce the risk of gout and hyperuricemia. Traditional bioenzyme based UA assays suffer from high cost, poor stability, inconvenience for storage and easy deactivation of bioenzymes. Wearable microfluidic colorimetric detection device for sweat UA detection has not been reported. The development of novel wearable microfluidic colorimetric detection chip with no requirement of bioenzymes for sweat UA detection is of great importance for health care monitoring. RESULTS: Firstly, Co@MnO2 nanozyme with high oxidase-like activity was synthesized and characterized. Co@MnO2 can catalyze the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) directly to generate blue-green colored ox-TMB. Green colored 2,2'-Azinobis-(3-ethylbenzthiazoline-6-sulphonate) radical (ABTS·+) was produced by the oxidation of ABTS by potassium persulfate. UA exhibits distinct quenching effect on Co@MnO2 catalyzed TMB colorimetric reaction system and ABTS·+ based colorimetric system, leading to obvious color fading of the two colorimetric systems. Then, a flexible microfluidic colorimetric detection chip for UA detection was fabricated by assembling Co@MnO2/TMB modified paper chips and ABTS·+ modified paper chips into a polydimethylsiloxane (PDMS) microfluidic chip. The fabricated microfluidic colorimetric detection chip exhibits good linear relationship for sweat UA detection. The linear range is from 20 to 200 µmol/L with detection limit as low as 6.6 µmol/L. Good results were obtained for the detection of UA in actual sweat from three volunteers. SIGNIFICANCE: This work provides two bio-enzyme free colorimetric detection systems for UA detection. Furthermore, a simple, low-cost and selective flexible wearable microfluidic colorimetric detection chip was fabricated for noninvasive and on-site detection of sweat UA, which holds great application potential for personal health monitoring and point-of-care testing.

NIR-II-Absorbing TMB Derivative for 1064 nm-Excited Photothermal Immunoassay.

Materials exhibiting strong absorption in the NIR-II region are appealing for photothermal conversion-based imaging, diagnosis, and therapy, due to better thermal effect and decreased absorption of water in such a region. 3,3',5,5'-Tetramethylbenzidine (TMB), the typical substrate in ELISA, has been explored in photothermal immunoassay, since its oxidation product (oxTMB) is photothermally active in the NIR region. However, its absorption at 1064 nm (the most often used laser wavelength in photothermal studies) is not appreciable, thus limiting the assay sensitivity. Here, we proposed a derivative of TMB (3,3'-dimethoxy-5,5'-dimethylbenzidine, 2-OCH3) bearing higher NIR-II absorption for 1064 nm-excited photothermal immunoassay. Since electron-donating groups can help decrease the energy gap of molecules (here -CH3 → -OCH3), the oxidation product of 2-OCH3 exhibited substantially red-shifted absorption as compared with oxTMB, leading to a more than twofold higher absorption coefficient at 1064 nm. As a result, 2-OCH3 showed enhanced sensitivity over TMB in a photothermal immunoassay (PTIA), yielding a limit of detection (LOD) of 0.1 ng/mL for prostate-specific antigen (PSA). The feasibility of 2-OCH3-based PTIA for diagnosis was further validated by analyzing PSA in 61 serum samples. Considering its superior photothermal performance, 2-OCH3 can be explored for a broad range of photothermal applications.

Single center utilization and post-transplant outcomes of thoracoabdominal normothermic regional perfusion deceased cardiac donor organs.

INTRODUCTION: Thoracoabdominal normothermic regional perfusion (TA-NRP) following cardiac death is an emerging multivisceral organ procurement technique. Recent national studies on outcomes of presumptive TA-NRP-procured organs are limited by potential misclassification since TA-NRP is not differentiated from donation after cardiac death (DCD) in registry data. METHODS: We studied 22 donors whose designees consented to TA-NRP and organ procurement performed at our institution between January 20, 2020 and July 3, 2022. We identified these donors in SRTR to describe organ utilization and recipient outcomes and compared them to recipients of traditional DCD (tDCD) and donation after brain death (DBD) organs during the same timeframe. RESULTS: All 22 donors progressed to cardiac arrest and underwent TA-NRP followed by heart, lung, kidney, and/or liver procurement. Median donor age was 41 years, 55% had anoxic brain injury, 45% were hypertensive, 0% were diabetic, and median kidney donor profile index was 40%. TA-NRP utilization was high across all organ types (88%-100%), with a higher percentage of kidneys procured via TA-NRP compared to tDCD (88% vs. 72%, p = .02). Recipient and graft survival ranged from 89% to 100% and were comparable to tDCD and DBD recipients (p ≥ .2). Delayed graft function was lower for kidneys procured from TA-NRP compared to tDCD donors (27% vs. 44%, p = .045). CONCLUSION: Procurement from TA-NRP donors yielded high organ utilization, with outcomes comparable to tDCD and DBD recipients across organ types. Further large-scale study of TA-NRP donors, facilitated by its capture in the national registry, will be critical to fully understand its impact as an organ procurement technique.

Nanozyme sensor array based on Fe, Se co-doped carbon material for the discrimination of Sulfur-containing compounds.

Developing methods for the accurate identification and analysis of sulfur-containing compounds (SCCs) is of great significance because of their essential roles in living organisms and the diagnosis of diseases. Herein, Se-doping improved oxidase-like activity of iron-based carbon material (Fe-Se/NC) was prepared and applied to construct a four-channel colorimetric sensor array for the detection and identification of SCCs (including biothiols and sulfur-containing metal salts). Fe-Se/NC can realize the chromogenic oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by activating O2 without relying on H2O2, which can be inhibited by different SCCs to diverse degrees to produce different colorimetric response changes as "fingerprints" on the sensor array. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) revealed that nine kinds of SCCs could be well discriminated. The sensor array was also applied for the detection of SCCs with a linear range of 1-50 µM and a limit of detection of 0.07-0.2 µM. Moreover, colorimetric sensor array inspired by the different levels of SCCs in real samples were used to discriminate cancer cells and food samples, demonstrating its potential application in the field of disease diagnosis and food monitoring. ENVIRONMENTAL IMPLICATIONS: In this work, a four-channel colorimetric sensor array for accurate SCCs identification and detection was successfully constructed. The colorimetric sensor array inspired by the different levels of SCCs in real samples were also used to discriminate cancer cells and food samples. Therefore, this Fe-Se/NC based sensor array is expected to be applied in the field of environmental monitoring and environment related disease diagnosis.

Nanozyme-induced deep learning-assisted smartphone integrated colorimetric and fluorometric dual-mode for detection of tetracycline analogs.

In this work, a colorimetric and fluorescent dual-mode probe controlled by NH2-MIL-88 B (Fe, Ni) nanozymes was developed to visually detect tetracycline antibiotics (TCs) residues quantitatively, as well as accurately distinguish the four most widely used tetracycline analogs (tetracycline (TC), chrycline (CTC), oxytetracycline (OTC), and doxycycline (DC)). Colorless substrate 3,3',5,5'-tetramethylbenzidine (TMB) may be oxidized to blue oxidized TMB by the Fe Fenton reaction, which was catalyzed by the NH2-MIL-88 B (Fe, Ni) nanozyme with POD-like activity. The colorimetric detection system allows TCs to interact with NH2-MIL-88 B (Fe, Ni). This inhibits the production of ·OH, weakens the oxidation process of TMB, and ultimately lightens the blue color in the system by blocking the electron transfer between NH2-MIL-88 B (Fe, Ni) and H2O2. Furthermore, TCs can interact with NH2-MIL-88 B (Fe, Ni) as a result of the internal filtering effect, which causes the fluorescence intensity to decrease as TCs concentration increases. Additionally, a portable instrument that combines a smartphone sensing platform with colorimetric and fluorescent signals was created for the quick, visual quantitative detection of TCs. The colorimetric and fluorescent dual-mode nano platform enables color change, with detection limits (LODs) of 0.182 µM and 0.0668 µM for the spectrometer and smartphone sensor, respectively, based on the inhibition of fluorescence and enzyme-like activities by TCs. Overall, the colorimetric and fluorescence dual-mode sensor has good stability, high specificity, and an efficient way to eliminate false-positive issues associated with a single detection mode.

CoO/Co-graphene quantum dots as an oxidative mimetic nanozyme for the colorimetric detection of L-cysteine.

The preparation of cobalt-based nanozymes with high oxidase-like activity still needs more efforts. In this paper, we report the synthesis of a CoO/Co-tryptophan-functional graphene quantum dot hybrid (CoO/Co-Try-GQD). Firstly, cobalt ions coordinate with the indole nitrogen on Try-GQD to form a complex, followed by thermal reduction and oxidation. The resulting hybrid presents a three-dimensional network structure, and CoO/Co nanoparticles are uniformly dispersed on the graphene sheet with an average size of 10 ± 0.24 nm. This unique structure improved the oxidase-like activity of the hybrid, enabling it to catalyze the oxidation of colorless 3,3',5,5'-tetramethylbenzidine (TMB) to rapidly produce deep blue ox-TMB with a strong absorbance at 652 nm (A652). A colorimetric method was developed for the highly sensitive determination of L-cysteine (L-cys) based on the inhibition of the hybrid's oxidase-like activity and low A652 caused by the binding of L-cys with Co atoms on CoO/Co via the Co-S bond. The A652 linearly decreased with increasing L-cys concentration in the range of 0.05-2 µM, and the detection limit was 0.032 µM. Further, the established method has been successfully applied to the determination of L-cys in milk.

MOF-polymer composites with well-distributed gold nanoparticles for visual monitoring of homocysteine.

The distribution of gold nanoparticles (AuNPs) on the surface of a metal-organic framework (MOF) plays a crucial role in the catalytic performance of MOF-AuNP composites. This study describes how the physical adsorption (PH@AuNPs-on-U) and chemical modification of AuNPs on the surface of UiO-66-NH2 (U) affect the composites' catalytic efficiency. After 2-vinyl-4,4-dimethyl-2-oxazolin-5-one (VD) linked to poly(N-2-hydroxypropyl methacrylamide) (PH) with U (UVD-PH), UVD-PH@AuNPs composites were constructed with PH as the capping and reducing reagent. The composites exhibited higher peroxidase (POD)-like activity than PH@AuNPs-on-U for oxidising 3,3'5,5'-tetramethylbenzidine (TMB) with H2O2. The approach demonstrated that the proposed composite-based nanozymes could significantly enhance their catalytic activity and had a highly uniform distribution of PH@AuNPs on the surface of UVD. An assay with the nanozymes for visual detection of homocysteine (Hcy) was developed, displaying a good linear relationship (R2 = 0.998) ranging from 3.34 µM to 30.0 µM and a detection of limit of 0.3 µM. Additionally, the UVD-PH@AuNPs-TMB-H2O2 system successfully monitored serum Hcy after intraperitoneal injection in rats. This study paves a new way for developing MOF-AuNPs with highly uniform surface distribution of polymer@AuNPs to boost its catalytic activity and to detect drugs in real bio-samples.

Mn-modified porphyrin metal-organic framework mediated colorimetric and photothermal dual-channel probe for sensitive detection of organophosphorus pesticides.

Herein, a novel dual-mode probe for organophosphorus pesticides (OPs) colorimetric and photothermal detection was developed based on manganese modified porphyrin metal-organic framework (PCN-224-Mn). PCN-224-Mn had excellent oxidase-like activity and oxidized colorless 3,3,5,5-tetramethylbenzidine (TMB) to blue-green oxidation state TMB (oxTMB), which exhibited high temperature under near-infrared irradiation. l-ascorbate-2-phosphate was hydrolyzed by acid phosphatase to produce ascorbic acid, which weakened colorimetric and photothermal signals by impacting oxTMB generation. The presence of OPs blocked the production of ascorbic acid by irreversibly inhibiting the activity of acid phosphatase, causing the restoration of chromogenic reaction and the increase of temperature. Under the optimal conditions, the probe showed a good linear response to OPs in the concentration range of 5 âˆ¼ 10000 ng/mL, using glyphosate as the analog. The detection limits of glyphosate in colorimetric mode and photothermal mode were 1.47 ng/mL and 2.00 ng/mL, respectively. The probe was successfully used for sensitive identification of OPs residues in tea, brown rice, and wheat flour. This work proposes a simple and reliable colorimetric/photothermal platform for OPs identification, which overcomes the problem that single-mode detection probes are susceptible to external factors, and has broad application potential in the field of food safety.

Hydrogel-involved portable colorimetric sensor based on oxidase mimic Fe/Co-NC for acetylcholinesterase detection and pesticides inhibition assessment.

Herein, we synthesized a novel N-doped carbon layer encapsulated Fe/Co bimetallic nanoparticles (Fe/Co-NC), which exhibited superior oxidase-like activity due to the facilitation of electron penetration and the formation of metal-nitrogen active sites. Fe/Co-NC could catalyze the oxidation of 3,3,5,5-tetramethylbenzidine (TMB) to blue oxTMB. Acetylcholinesterase (AChE) could catalyze the hydrolysis of thioacetylcholine to produce reducing thiocholine, which prevented TMB from oxidation. Thus, a portable hydrogel colorimetric sensor was developed for on-site and visual monitoring of AChE with the detection limit of 0.36 U L-1, and successfully applied to detect AChE in human erythrocyte samples. Furthermore, this platform was used to investigate the inhibition of triazophos on AChE activity.

Colorimetric and electrochemical dual-mode uric acid determination utilizing peroxidase-mimicking activity of CoCu bimetallic nanoclusters.

We present the preparation of CoCu bimetallic nanoclusters (Co@Cu-BNCs) by a hydrothermal and one-step pyrolysis method to build a colorimetric and electrochemical dual-mode sensing platform for uric acid (UA) detection. In the presence of H2O2, Co@Cu-BNCs with peroxidase-mimicking activity may convert colorless 3,3',5,5'-tetramethylbenzidine (TMB) to blue-colored oxidized TMB (oxTMB). However, due to the inhibitory effect of uric acid (UA) on the oxidation process of TMB, the characteristic absorption peak intensity of oxTMB decreased when UA was added into a mixed solution. In this approach, a colorimetric assay platform for the detection of UA was demonstrated, with a linear range of 0.1-195 µM and a low limit of detection of 0.06 µM (S/N ratio of 3). In addition, an even wider detection range is achieved in the electrochemical method, due to the pronounced electrocatalytic activity of Co@Cu-BNCs. The surface of the glassy carbon electrode was modified with Co@Cu-BNCs to build an electrochemical sensor for detecting UA. The sensor achieves a wider linear range from 2 to 1000 µM and a limit of detection of 0.61 µM (S/N ratio of 3). Moreover, the detection of UA in a human serum sample showed satisfactory results. The results proved that the colorimetric and electrochemical dual-mode detection platform was sensitive, convenient and accurate.

A portable intelligent hydrogel platform for multicolor visual detection of HAase.

Hyaluronidase (HAase) is an important endoglycosidase involved in numerous physiological and pathological processes, such as apoptosis, senescence, and cancer progression. Simple, convenient, and sensitive detection of HAase is important for clinical diagnosis. Herein, an easy-to-operate multicolor visual sensing strategy was developed for HAase determination. The proposed sensor was composed of an enzyme-responsive hydrogel and a nanochromogenic system (gold nanobipyramids (AuNBPs)). The enzyme-responsive hydrogel, formed by polyethyleneimine-hyaluronic acid (PEI-HA), was specifically hydrolyzed with HAase, leading to the release of platinum nanoparticles (PtNPs). Subsequently, PtNPs catalyzed the mixed system of 3,3',5,5'-tetramethylbenzidine (TMB) and H2O2 to produce TMB2+ under acidic conditions. Then, TMB2+ effectively etched the AuNBPs and resulted in morphological changes in the AuNBPs, accompanied by a blueshift in the localized surface plasmon resonance peak and vibrant colors. Therefore, HAase can be semiquantitatively determined by directly observing the color change of AuNBPs with the naked eye. On the basis of this, the method has a linear detection range of HAase concentrations between 0.6 and 40 U/mL, with a detection limit of 0.3 U/mL. In addition, our designed multicolor biosensor successfully detected the concentration of HAase in human serum samples. The results showed no obvious difference between this method and enzyme-linked immunosorbent assay, indicating the good accuracy and usability of the suggested method.

Marine bromophenols suppressed choroidal neovascularization by targeting HUWE1 through NF-κb signaling pathway.

Inflammation plays a key role in the progression of choroidal neovascularization (CNV). Regular intravitreal injection of anti-VEGF medication is required for many patients to sustain eye condition as CNV always recurs due to persistent chronic inflammation in the retina and choroid. Marine bromophenols (BDB) have been widely studied due to their diverse bioactivities, including anti-inflammatory effect, though the mechanism of which remained unclear. Our study demonstrated that BDB could restricted endothelial cells' function and suppressed choroidal explants both in vitro and in vivo without out affecting the cells viability. BDB also significantly reduced numerous inflammatory cytokines in both raw cells and choroidal tissue, including IL-1ß, IL-6, TNF-α, IL-4 and MMP-9. Moreover, we demonstrated that BDB down regulated phosphorylation of NF-κB p65 in the raw cells. By Co-IP assay, HUWE1 was found to be bound with BDB and the binding location was at sequences position 4214. When overexpressed HUWE1 in HUVECs, the suppression of endothelial cells' function by BDB became more significant. Taken together, the findings in this study showed that BDB suppressed endothelial cells' function and choroidal neovascularization by targeting HUWE1 through NF-κB pathway, which suggested that BDB could be a potential therapeutic candidate in treating chronic inflammation in choroidal neovascularization.

Decoupling of the Confused Complex in Oxidation of 3,3',5,5'-Tetramethylbenzidine for the Reliable Chromogenic Bioassay.

Regulation of the reaction pathways is a perennial theme in the field of chemistry. As a typical chromogenic substrate, 3,3',5,5'-tetramethylbenzidine (TMB) generally undertakes one-electron oxidation, but the product (TMBox1) is essentially a confused complex and is unstable, which significantly hampers the clinic chromogenic bioassays for more than 50 years. Herein, we report that sodium dodecyl sulfate (SDS)-based micelles could drive the direct two-electron oxidation of TMB to the final stable TMBox2. Rather than activation of H2O2 oxidant in the one-electron TMB oxidation by common natural peroxidase, activation of the TMB substrate by SDS micelles decoupled the thermodynamically favorable complex between TMBox2 with unreacted TMB, leading to an unusual direct two-electron oxidation pathway. Mechanism studies demonstrated that the complementary spatial and electrostatic isolation effects, caused by the confined hydrophobic cavities and negatively charged outer surfaces of SDS micelles, were crucial. Further cascading with glucose oxidase, as a proof-of-concept application, allowed glucose to be more reliably measured, even in a broader range of concentrations without any conventional strong acid termination.

N-Rich and Sulfur-Doped Nano Hollow Carbons with High Oxidase-like Activity Prepared Using a Green Template of CaCO3 for Bacteriostasis.

Nanozymes, enzyme-mimicking nanomaterials, have attracted increasing attention due to their low cost, high stability, and catalytic ability compared with natural enzymes. However, the catalytic efficiency of the nanozymes is still relatively low, and catalytic reaction mechanisms remain unclear. To address these issues, herein we prepared nitrogen-riched and sulfur-codoped nano hollow carbons (N/S-HCS) using a green and useful template of CaCO3. N/S-HCS exhibits enhanced oxidase-like activity and catalytic kinetic performance. It could directly oxidize the colorless 3,3',5,5'-tetramethylbenzidine (TMB) to the heavy blue colored ox-TMB without H2O2. The maximum reaction rate (Vmax) is 186.7 × 10-8 M·s-1, and Michaelis-Menten constant (Km) is 0.162 mM. DFT results show that N and S codoping could work synergistically to provide more active sites, resulting in the superior ability to adsorb oxygen and enhanced catalytic activity. Meantime, we develop a multispectral characterization strategy to unravel catalytic reaction mechanisms about N/S-HCS. It successfully induces the generation of superoxide (•O2-) and hydroxyl (•OH) during the colorimetric reaction which are the key intermediate products of the catalytic reaction. Furthermore, N/S-HCS increased the cellular reactive oxygen species level significantly and induced bacteriostasis to more than 95% of Escherichia coli.

Impact of bridge heterology on functional parameters of ELISA for 17α-methyltestosterone.

In this study, we have developed bridge heterologous ELISA for the detection of 17α- Methyltestosterone by incorporating aromatic spacers between 17α-Methyltestosterone-3-Carboxymethyloxime and Horseradish peroxidase label through N-hydroxysuccinimide mediated carbodiimide reaction method. The immunogen 17α-Methyltestosterone-3-Carboxymethyloxime-Bovine serum albumin used to generate the antibody was also prepared by the N-hydroxysuccinimide mediated carbodiimide reaction without using any spacer. We have studied the impact of bridge/aromatic spacers on functional parameters i.e. sensitivity, affinity and ED50 of the bridge heterologous assay and compared it with homologous assay. The five combinations of bridge heterologous assay using 17α-Methyl testosterone-3-CMO-BSA antiserum and 17α-MT-3-CMO-4,4'-Diaminodiphenyl sulphide-HRP, 17α MT-3-CMO-4,4'-Oxydianiline-HRP, 17α-MT-3-CMO-Benzidine-HRP, 17α- MT-3-CMO-p-Phenylenediamine-HRP and 17α-MT-3-CMO-Dapson-HRP enzyme conjugates were evaluated. Out of these five combinations, the combination 17α-MT-3-CMO-BSA with 17α-MT-3-CMO-Benzidine-HRP showed the best results. Sensitivity, affinity and ED50 were improved and found to be 0.02 ng/mL, 0.086 × 10-8 L/mol and 2.95 ng/mL than homologous assay where Sensitivity, affinity and ED50 were 0.11 ng/mL, 0.02 × 10-8 L/mol and 5.78 ng/mL respectively. The cross-reactivity for this bridge heterologous assay combination was seen with only 4 steroids (6-hydrotestosterone- 6%, Testosterone-5.14%, Danazol-0.9% and Nandrolone-0.85%) instead of eight steroids (6-hydrotestosterone-43.75%, Testosterone-38.3%, Danazol-25.14%, Androstenediol-19.16%, Nandrolone-19%, Metandienone-5%, Androstenedione-3.52%, and 17α dimethyltestosterone-2%) as in homologous assay out of 59 structurally related steroids. Thus, the results of this study conclude that the incorporation of aromatic spacer (bridge) in enzyme conjugate has a crucial role in improving sensitivity, specificity, ED50 and affinity of the developed assay. The assay was then studied for parameters such as recovery (97.4%-108.6%), precision (Inter and Intra-assay coefficient of variation <10%), correlation coefficient (R2 = 0.96) by comparing with the commercial kit and validated by measuring levels of 17α- methyltestosterone in rat serum after administering them.

Nanozyme based colorimetric detection of biogenic gaseous H2S using Ag@Au core/shell nanoplates with peroxidase-like activity.

A highly sensitive and facile colorimetric assay is introduced for detecting biogenic gaseous H2S using peroxidase (POD)-like catalytic activity of silver core/gold shell nanoplates (Ag@Au NPls). H2S can react with Ag@Au NPls to form Ag2S or Au2S on their surface, which can reduce POD-like activity of Ag@Au NPls and consequently decrease the absorbance at 650 nm due to oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of hydrogen peroxide (H2O2). For in situ and multiple detection of H2S, we utilized a microplate cover with 24 polydimethylsiloxane inner wells where Ag@Au NPls reacted with H2S gas followed by treatment with TMB/H2O2. As a result, the change in absorbance at 650 nm showed a linear relationship with the H2S concentration in the range 0.33 to 2.96 µM (0.36 absorbance/µM H2S in PBS, R2 = 0.994) with a limit of detection of 263 nM and a relative standard deviation of 4.4%. Finally, this assay could detect H2S released from Eikenella corrodens, used as a model bacterium, in a short time (20 min) or at a low number of bacteria (1 × 104 colony forming units/mL). Therefore, this assay is expected to be applied for the study of H2S signaling in bacterial physiology, as well as measure H2S production released from other oral bacteria that cause halitosis and oral diseases, leading to the subsequent diagnosis.

Enhancement of the peroxidase-like activity of hollow spherical FexNi1-xS2/SC nanozymes.

Artificial nanozymes have been receiving considerable interest for their outstanding performance and wide application. However, their low activity results in a high concentration of substrates, costs, and environmental pollution. To enhance nanozymic activity, a composite, FexNi1-xS2/hollow carbon spheres (FexNi1-xS2/SC), was facilely synthesized by a solvothermal method. The response surface methodology (RSM) was used to optimize the Ni content in FexNi1-xS2/SC and the experimental conditions, where Fe0.75Ni0.25S2/SC exhibited the highest activity. The Km (Michaelis-Menten's constant) values of Fe0.75Ni0.25S2/SC are 0.025 and 0.021 mM with H2O2 and oxidized 3,3',5,5'-tetramethylbenzidine (TMB) as the substrates, respectively, which are 148 times and 20.5 times lower than those with HRP, 1.88 and 7.19 times lower than those of FeS2/SC, and 1.88 and 10.52 times lower than those of Fe0.8Ni0.2S2, meaning a strong affinity of Fe0.75Ni0.25S2/SC for the substrate. The catalytic efficiency (Kcat/Km) of Fe0.75Ni0.25S2/SC was 5.4 (H2O2) and 27.4 times (TMB), and 9.7 (H2O2) and 66.2 times (TMB) higher than those of FeS2/SC and Fe0.8Ni0.2S2, respectively. The effects of the synergistic interaction between Fe and Ni, the S-C bond formation, and the hollow carbon spheres on the activity were studied. A nanozymic mechanism was proposed. Fe0.75Ni0.25S2/SC could be used to detect cysteine (Cys) at room temperature in 1 min with a detection limit (LOD) of 0.049 µM.

Diazo-reaction based dual-mode colorimetric-electrochemical sensing of nitrite in pickled food.

Development of an effective and convenient sensor for sensitive detection of nitrites is of great concern since excessive amounts of nitrites can be harmful to both human health and the environment. In this work, Cu-MOF modified exfoliated graphite paper (EGP) was employed as a signal reporter to enable the visual and electrochemical dual-mode sensing of nitrites. Cu-MOFs were in situ synthesized on EGP, which exhibited an excellent oxidase enzyme-like activity to oxidize 3,3',5,5'-tetramethylbenzidine (TMB) into its oxidation product (oxTMB). The multi-layer structure and the superior electrical conductivity of EGP not only facilitated the loading of the Cu-MOF nanozyme for colorimetric sensing but also enabled its use as an underlying backbone to support electroanalysis. Based on the recognition of nitrite through a highly specific diazo reaction between nitrite and oxTMB, the addition of nitrite caused the colorimetric sensing solution to change color from blue to green, which allowed for the colorimetric sensing of nitrite with a limit of detection (LOD) of 8.5 × 10-6 mol L-1. Meanwhile, the Cu-MOF/EGP electrochemical platform was employed for ratiometric detection of nitrite based on the electrochemical oxidation of nitrite and TMB. Compared with the colorimetric mode, the electrochemical mode possessed higher sensitivity with a LOD of 5.4 × 10-7 mol L-1, indicating the high sensitivity and accuracy of the proposed dual-mode sensing strategy. Furthermore, the determination of nitrite in different pickled food samples is demonstrated.