Employing the Anchor DSPE-PEG as a Redox Probe for Ratiometric Electrochemical Detection of Surface Proteins on Extracellular Vesicles with Aptamers.

Quantitative analysis of surface proteins on extracellular vesicles (EVs) has been considered to be a crucial approach for reflecting the status of diseases. Due to the diverse composition of surface proteins on EVs and the interference from nonvesicular proteins, accurately detecting the expression of surface proteins on EVs remains a challenging task. While membrane affinity molecules have been widely employed as EVs capture probes to address this issue, their inherent biochemical properties have not been effectively harnessed. In this paper, we found that the electrochemical redox activity of the DSPE-PEG molecule was diminished upon its insertion into the membrane of EVs. This observation establishes the DSPE-PEG molecule modified on the Au electrode surface as a capture and a redox probe for the electrochemical detection of EVs. By utilizing methylene blue-labeled aptamers, the targeted surface proteins of EVs can be detected by recording the ratio of the oxidation peak current of methylene blue and DSPE-PEG. Without complicated signal amplification, the detection limit for EVs is calculated to be 8.11 × 102 particles/mL. Using this platform, we directly analyzed the expression of CD63 and HER2 proteins on the surface of EVs in human clinical plasma samples, demonstrating its significant potential in distinguishing breast cancer patients from healthy individuals.

Correction: 'Redox-mediated carbon monoxide release from a manganese carbonyl-implications for physiological CO delivery by CO releasing moieties' (2021), by Barrett et al.

[This corrects the article DOI: 10.1098/rsos.211022.][This corrects the article DOI: 10.1098/rsos.211022.].

Preparation and investigation of novel endopeptidase-exopeptidase co-immobilized nanoflowers with improved cascade hydrolysis.

Enzymatic hydrolysis is a promising approach for protein and food processing. However, the efficiency of this approach is constrained by the self-hydrolysis, self-agglomeration of free enzymes and the limited applicability resulted from enzymes' selectivityt. Here, novel organic-inorganic hybrid nanoflowers (AY-10@AXH-HNFs) were prepared by coordinating Cu2+ with both endopeptidase of PROTIN SD-AY10 and exopeptidase of Prote AXH. The results indicate that the AY-10@AXH-HNFs exhibited 4.1 and 9.6 times higher catalytic activity than free Prote AXH and PROTIN SD-AY10, respectively, for the enzymatic hydrolysis of N-benzoyl-L-arginine ethyl ester (BAEE). The kinetic parameters of Km, Vmax and Kcat/Km by AY-10@AXH-HNFs were determined to be 0.6 mg/mL, 6.8 mL·min/mg and 6.1 mL/(min·mg), respectively, surpassing the values obtained from free endopeptidase and exopeptidase. Furthermore, the ability of AY-10@AXH-HNFs to retain 41 % of their initial catalytic activity after undergoing 5 cycles of repeated use confirmed their stability and reusability. This study introduces a novel approach of co-immobilizing endopeptidase and exopeptidase on nanoflowers, resulting in significantly enhanced stability and reusability of the protease in catalytic applications.

Fabrication of a Disposable Amperometric Sensor for the Determination of Nitrite in Food.

Silver nanoparticles (AgNPs) were synthesized through an environmentally friendly method with tea extract as a reduction agent. By immobilizing them on the surface of a low-cost pencil graphite electrode (PGE) with the aid of a simple and well-controlled in-situ electropolymerization method, a novel nanosensing interface for nitrite was constructed. The film-modified PGE showed good electrocatalytic effects on the oxidation of nitrite and was characterized through scanning electron microscopy, X-ray photoelectron spectroscopy, and electrochemical techniques. Characterization results clearly show that the successful modification of AgNPs improved the surface area and conductivity of PGEs, which is beneficial to the high sensitivity and short response time of the nitrite sensor. Under the optimal detection conditions, the oxidation peak current of nitrite had a good linear relationship with its concentration in the range of 0.02-1160 µmol/L with a detection limit of 4 nmol/L and a response time of 2 s. Moreover, the sensor had high sensitivity, a wide linear range, a good anti-interference capability, and stability and reproducibility. Additionally, it can be used for the determination of nitrite in food.

Cu-In-S/ZnS:Gd3+ quantum dots with isolated fluorescent and paramagnetic modules for dual-modality imaging in vivo.

Gd3+-doped quantum dots (QDs) have been widely used as small-sized bifunctional contrast agents for fluorescence/magnetic resonance (FL/MR) dual-modality imaging. However, Gd3+ doping will always compromise the FL of host QDs. Therefore, balancing the Gd3+ doping and the optical properties of QDs is crucial for constructing high-performance bifunctional nanoprobes. Additionally, most paramagnetic QDs are synthesized in the organic phase and need to be transferred to the aqueous phase for bioimaging. Herein, ingeniously designed shell-doped Cu-In-S/ZnS:Gd3+ QDs have been prepared in the aqueous phase. It has been demonstrated that isolating paramagnetic Gd3+ from fluorescent Cu-In-S core via doping Gd3+ into ZnS shell not only avoided the decrease of FL quantum yield (QY), but also ensured the water accessibility of paramagnetic Gd3+ ions, by which the FL QY and r1 relaxivity of Cu-In-S/ZnS:Gd3+ QDs achieved as much as 15.6% and 15.33 mM-1·s-1, respectively. These high-performance QDs with excellent stability, low biotoxicity, and good tumor permeability were successfully applied for in vivo tumor FL/MR dual-modality imaging, and have shown significant potential in the precision detection and diagnosis of diseases.

An Amperometric Biomedical Sensor for the Determination of Homocysteine Using Gold Nanoparticles and Acetylene Black-Dihexadecyl Phosphate-Modified Glassy Carbon Electrode.

A novel nanocomposite film composed of gold nanoparticles and acetylene black-dihexadecyl phosphate was fabricated and modified on the surface of a glassy carbon electrode through a simple and controllable dropping and electropolymerization method. The nanocomposite film electrode showed a good electrocatalytic response to the oxidation of homocysteine and can work as an amperometric biomedical sensor for homocysteine. With the aid of scanning electron microscopy, energy dispersive X-ray technology and electrochemical impedance spectroscopy, the sensing interface was characterized, and the sensing mechanism was discussed. Under optimal conditions, the oxidation peak current of homocysteine was linearly increased with its concentration in the range of 3.0 µmol/L~1.0 mmol/L, and a sensitivity of 18 nA/(µmol/L) was obtained. Furthermore, the detection limit was determined as 0.6 µmol/L, and the response time was detected as 3 s. Applying the nanocomposite film electrode for monitoring the homocysteine in human blood serum, the results were satisfactory.

Reversible covalent nanoassemblies for augmented nuclear drug translocation in drug resistance tumor.

The drug efflux by P-glycoprotein (P-gp) is the primary contributor of multidrug resistance (MDR), which eventually generates insufficient nuclear drug accumulation and chemotherapy failure. In this paper, reversible covalent nanoassemblies on the basis of catechol-functionalized methoxy poly (ethylene glycol) (mPEG-dop) and phenylboronic acid-modified cholesterol (Chol-PBA) are successfully synthesized for delivery of both doxorubicin (DOX, anti-cancer drug) and tariquidar (TQR, P-glycoprotein inhibitor), which shows efficient nuclear DOX accumulation for overcoming tumor MDR. Through naturally forming phenylboronate linkage in physiological circumstances, Chol-PBA is able to bond with mPEG-dop. The resulting conjugates (PC) could self-assemble into reversible covalent nanoassemblies by dialysis method, and transmission electron microscopy analysis reveals the PC distributes in nano-scaled spherical particles before and after drug encapsulation. Under the assistance of Chol, PC can enter into lysosome of tumor cells via low-density lipoprotein (LDL) receptor-mediated endocytosis. Then the loaded TQR and DOX are released in acidic lysosomal compartments, which inhibit P-gp mediated efflux and elevate nuclear accumulation of DOX, respectively. At last, this drug loaded PC nanoassemblies show significant tumor suppression efficacy in multidrug-resistant tumor models, which suggests great potential for addressing MDR in cancer therapy.

An improved automated diatom detection method based on YOLOv5 framework and its preliminary study for taxonomy recognition in the forensic diatom test.

The diatom test is a forensic technique that can provide supportive evidence in the diagnosis of drowning but requires the laborious observation and counting of diatoms using a microscopy with too much effort, and therefore it is promising to introduce artificial intelligence (AI) to make the test process automatic. In this article, we propose an artificial intelligence solution based on the YOLOv5 framework for the automatic detection and recognition of the diatom genera. To evaluate the performance of this AI solution in different scenarios, we collected five lab-grown diatom genera and samples of some organic tissues from drowning cases to investigate the potential upper/lower limits of the capability in detecting the diatoms and recognizing their genera. Based on the study of the article, a recall score of 0.95 together with the corresponding precision score of 0.9 were achieved on the samples of the five lab-grown diatom genera via cross-validation, and the accuracy of the evaluation in the cases of kidney and liver is above 0.85 based on the precision and recall scores, which demonstrate the effectiveness of the AI solution to be used in drowning forensic routine.

Pathway of Diatoms Enter Experimental Rabbits through the Lymphatic System of the Digestive Tract.

OBJECTIVES: To study whether diatoms can enter the body through the lymphatic system of the digestive tract. METHODS: Twenty experimental rabbits were divided into the test group and the control group randomly, and intragastric administration was performed with 20 mL water sample from the Pearl River and 20 mL ultrapure water, respectively. After 30 min, lymph, lungs, livers and kidneys were extracted for the diatom test. The concentration, size and type of diatoms were recorded. RESULTS: The concentration of diatoms of the test group was higher than that of the control group (P<0.05). In the test group, Stephanodiscus, Coscinodiscus, Cyclotella, Melosira, Nitzschia, Synedra, Cymbella, and Navicula were detected; in the control group, Stephanodiscus, Coscinodiscus and Cyclotella were detected. The long diameter and the short diameter of diatoms of the test group were higher than those of the control group (P<0.05). In the test group, 1-2 diatoms were detected in 3 lung samples and 2 liver samples, which were Stephanodiscus or Cyclotella, and no diatoms were detected in the kidney samples; in the control group, 1-2 diatoms were detected in 2 lung samples and 3 liver samples, which were Stephanodiscus or Coscinodiscus, and no diatoms were detected in the kidney samples. CONCLUSIONS: Diatoms can enter the body through the lymphatic fluid, which is one of the reasons for the presence of diatoms in tissues and organs of non-drowning cadavers.

Effects of Digestive Temperature and Time on Diatom Test.

OBJECTIVES: To study the effects of temperature and time for diatoms digestion and find out suitable digestive temperature and time. METHODS: Eighty pieces of liver tissues were collected, each piece of tissue was 2 g, and 2 mL Pearl River water was added to each piece of tissue. The digestion temperature was set at 100 ℃, 120 ℃, 140 ℃, 160 ℃, 180 ℃ and the digestion time was set at 40, 50, 60, 70, 80 min. The liver tissue and water mixture were divided into 8 portions in each group. All the samples were tested by microwave digestive - vacuum filtration - automated scanning electron microscopy method. The quantity of diatom recovered and the quality of residue on the membrane were recorded. RESULTS: When the digestion time was set to 60 min, there were statistically significant differences in the number of diatoms recovered at different temperatures (P<0.05). The maximum number of diatoms recovered was (28 797.50±6 009.67) at 140 ℃, and the minimum residue was (0.60±0.28) mg at 180 ℃. When the digestion temperature was set at 140 ℃, there were statistically significant differences in the number of diatoms recovered at different digestion times (P<0.05). The number of diatoms recovered was the highest at 40 min, it was up to (20 650.88±1 950.29), and the residue quality of each group had no statistical significance among different digestion time groups(P>0.05). CONCLUSIONS: The effect of diatom digestion is related to temperature and time. When the digestion temperature was 140 ℃ and the digestion time was 40, 50 and 60 min, it is favorable for diatom test.

Application of Diatoms Quantitative Analysis in the Diagnosis of Drowning.

OBJECTIVES: To retrospectively analyze diatom test cases of corpses in water and discuss the value of quantitative analysis of diatoms in the diagnosis of drowning. METHODS: A total of 490 cases of water-related death were collected. They were divided into drowning group and postmortem immersion group according to the cause of death. Diatoms in lung, liver, kidney tissue and water sample were analyzed quantitatively by microwave digestion-vacuum filtration-automated scanning electron microscopy (MD-VF-Auto SEM) method. The ratios of content of diatoms in lung tissue and water sample (CL/CD) were calculated. RESULTS: The results of diatom test for three organs (lung, liver and kidney) were all positive in 400 cases (85.5%); the content of diatom in lung, liver, kidney tissues, and water samples of drowning group were (113 235.9±317 868.1), (26.7±75.6), (23.3±52.2) and (12 113.3±21 760.0) cells/10 g, respectively; the species of diatom were (7.5±2.8), (2.6±1.9), (2.9±2.1) and (8.9±3.0) types, respectively; the CL/CD of drowning group and postmortem immersion group were (100.6±830.7) and (0.3±0.4), respectively. CONCLUSIONS: Quantitative analysis of diatoms can provide supportive evidence for the diagnosis of drowning, and the parameter CL/CD can be introduced into the analysis to make a more accurate diagnosis of drowning.

Aptamers as Recognition Elements for Electrochemical Detection of Exosomes.

Exosome analysis is emerging as an attractive noninvasive approach for disease diagnosis and treatment monitoring in the field of liquid biopsy. Aptamer is considered as a promising molecular probe for exosomes detection because of the high binding affinity, remarkable specificity, and low cost. Recently, many approaches have been developed to further improve the performance of electrochemical aptamer based(E-AB) sensors with a lower limit of detection. In this review, we focus on the development of using aptamer as a specific recognition element for exosomes detection in electrochemical sensors. We first introduce recent advances in evolving aptamers against exosomes. Then, we review methods of immobilization aptamers on electrode surfaces, followed by a summary of the main strategies of signal amplification. Finally, we present the insights of the challenges and future directions of E-AB sensors for exosomes analysis.

Concordance analysis of diatom types and patterns in lung tissue and drowning medium in laboratory animal model.

Diatom test has been widely used in the diagnosis of drowning and inferring the drowning site. One of the issues is whether the concordance of the diatom types and patterns between the drowning victim's organs and media should be considered an essential requirement for the diagnosis of drowning. In this study, lung tissues from 20 rabbits and drowning media were studied by the Microwave Digestion-Vacuum Filtration-Automated Scanning Electron Microscopy method, and four methods, type consistency, coefficient of similarity, squared-chord distance, and cluster analysis, were introduced to analyze the diatom types and patterns for evaluating the value of diatom consistency in drowning cases. The results showed that diatom types and patterns in lung tissues do not perfectly match the drowning medium, and they are sometimes concordant with the drowning medium sampled from other than drowning site. We should be cautious when using diatom detection to infer drowning sites.

Redox-mediated carbon monoxide release from a manganese carbonyl-implications for physiological CO delivery by CO releasing moieties.

The dynamics of hydrogen peroxide reactions with metal carbonyls have received little attention. Given reports that therapeutic levels of carbon monoxide are released in hypoxic tumour cells upon manganese carbonyls reactions with endogenous H2O2, it is critical to assess the underlying CO release mechanism(s). In this context, a quantitative mechanistic investigation of the H2O2 oxidation of the water-soluble model complex fac-[Mn(CO)3(Br)(bpCO2)]2-, (A, bpCO2 2- = 2,2'-bipyridine-4,4'-dicarboxylate dianion) was undertaken under physiologically relevant conditions. Characterizing such pathways is essential to evaluating the viability of redox-mediated CO release as an anti-cancer strategy. The present experimental studies demonstrate that approximately 2.5 equivalents of CO are released upon H2O2 oxidation of A via pH-dependent kinetics that are first-order both in [A] and in [H2O2]. Density functional calculations were used to evaluate the key intermediates in the proposed reaction mechanisms. These pathways are discussed in terms of their relevance to physiological CO delivery by carbon monoxide releasing moieties.

Automated diatom searching in the digital scanning electron microscopy images of drowning cases using the deep neural networks.

Forensic diatom test has been widely accepted as a way of providing supportive evidences in the diagnosis of drowning. The current workflow is primarily based on the observation of diatoms by forensic pathologists under a microscopy, and this process can be very time-consuming. In this paper, we demonstrate a deep learning-based approach for automatically searching diatoms in scanning electron microscopic images. Cross-validation studies were performed to evaluate the influence of magnification on performance. Moreover, various training strategies were tested to improve the performance of detection. The conclusion shows that our approach can satisfy the necessary requirements to be integrated as part of an automatic forensic diatom test.

Small-sized copolymeric nanoparticles for tumor penetration and intracellular drug release.

Poor solid-tumor penetration of nanocarriers limits the drug efficacy. Herein, small-sized copolymeric nanoparticles are prepared for delivering the chemotherapeutic drug DOX into solid tumors deeply and releasing the drug effectively. These small-sized copolymeric nanoparticles represent substantial potential for clinical translation.

A nanocomposite consisting of etched multiwalled carbon nanotubes, amino-modified metal-organic framework UiO-66 and polyaniline for preconcentration of polycyclic aromatic hydrocarbons prior to their determination by HPLC.

A polyaniline composite doped with etched multi-walled carbon nanotubes and UiO-66-NH2 was prepared by electropolymerization. It was used as a sorbent to extract the polycyclic aromatic hydrocarbons (PAHs) phenanthrene, fluoranthene and pyrene. Its surface morphology, crystal structure and capability of adsorbing PAHs were characterized by scanning electron microscopy, X-ray photoelectron spectrometry, Fourier transform infrared spectrometry and zeta potentiometry. The π stacking and anion-π interactions are shown to play dominant roles in the sorption mechanism. Coupled with high performance liquid chromatography, the composite-modified fiber was applied to detect PAHs in lake water samples by direct immersion extraction. The method excels by (a) wide linear range (0.05-20 ng mL-1), (b) low limits of detection (10 pg mL-1), (c) satisfactory recovery from spiked samples (84.7-113.8%), and (d) good reproducibility (relative standard deviations of <6.5%). The method is superior in terms of costs and reproducibility compared to some pretreatment methods with mass spectrometric detection. Graphical abstractSchematic representation for interaction between PANI-etched MWCNT/UiO-66-NH2 and polycyclic aromatic hydrocarbons (phenanthrene, fluoranthene, pyrene).

Evaluation of L/D ratio in a water-related case for the differentiation between drowning and postmortem immersion.

It is a difficult task to determine the cause of death for decomposed bodies found in water. Diatom test has been shown to provide supportive evidence for the diagnosis of drowning. According to our previous studies, the ratio of diatom content in lung tissue to diatom content in drowning medium (L/D ratio) is a very useful indicator to distinguish between drowning and postmortem immersion. In this article, we presented a case of a highly decomposed body being recovered from water with no significant findings on its cause of death. We applied the microwave digestion-vacuum filtration-automated scanning electron microscopy method (MD-VF-Auto SEM) to detect diatoms in the organs and suspected drowning medium. In this case, positive results are found in diatom tests of lung tissue, liver tissue and kidney tissue, while the L/D ratio analysis suggested that the victim was more likely to suffer from postmortem immersion. Taking the result of L/R ratio and other findings into consideration, we can eliminate the possibility of drowning. With continuous study, the L/D ratio analysis would be a valuable tool in diagnosis of drowning.

Detection of theophylline using molecularly imprinted mesoporous silica spheres.

Here, we report a three-dimensional (3D) network molecularly imprinted polymer (MIP) on electrode surface to achieve an efficient and specific detection of theophylline in foodstuffs, using theophylline as the template molecule, mesoporous silica nanospheres (MSNs) as the signal transducer to shuttle electrons, and both phenyltriethoxysilane and pyrrole as functional monomers. The electron microscope images reveal the presence of well-distributed hierarchically MSNs with a pomegranate-like morphology, topped with MIP uniform layer. Electrochemical characterizations were carried out to monitor the properties of the resulting sensing platform based on the MIP/gate effect employing hexacyanoferrate molecules as the electrochemical probe. The data show that due to the high conductivity and electron transfer ability of the prepared theophylline-imprinted membrane, this method exhibits excellent sensitivity and binding affinity with a linear dynamic concentration range in excess of six orders of magnitude and low detection limit (0.66 nM), meeting the requirements of theophylline trace analysis.

Graphene oxide reinforced core-shell structured Ag@Cu2O with tunable hierarchical morphologies and their morphology-dependent electrocatalytic properties for bio-sensing applications.

In this study, a facile solution approach was developed for the synthesis of a series of core-shell structured Ag@Cu2O nanocrystals of various shapes including triangles, spheres, and cubes with well-defined stable heterojunctions. The electrooxidation of dopamine (DA), uric acid (UA), guanine (G), and adenine (A) using these hybrids revealed morphology-dependent sensing properties, with activities and accumulation ability following the order, triangular Ag@Cu2O > spherical Ag@Cu2O > cubic Ag@Cu2O. Further, we constructed a novel graphene oxide (GO) nanosheet-reinforced triangular Ag@Cu2O ternary hetero-nanostructure. Such a hybrid with a three-dimensional interconnected hierarchical architecture is suitable for catalysis, since it not only leads to improved interfacial electron transfer, but also readily exposes the highly catalytic Ag@Cu2O to the reactants. Therefore, more enhanced electrochemical activities were observed for the oxidation of DA, UA, G, and A. This study provides an efficient way to synthesize morphology-controlled Ag@Cu2O heterogeneous catalysts for the fabrication of potential biosensors, and also opens up attractive avenues in the design of multifunctional ternary noble metal-semiconductor-carbon hybrids.