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Tumor-derived extracellular vesicles (EVs) carry tumor-specific proteins and RNAs, thus becoming prevalent targets for early cancer diagnosis. However, low expression of EV cargos and insufficient diagnostic power of individual biomarkers hindered EVs application in clinical practice. Herein, we propose a multiplex Codetection platform of proteins and RNAs (Co-PAR) for EVs. Co-PAR adopted a pair of antibody-DNA probes to recognize the same target protein, which in turn formed a double-stranded DNA. Thus, the target protein could be quantified by detecting the double-stranded DNA via qPCR. Meanwhile, qRT-PCR simultaneously quantified the target RNAs. Thus, with a regular qPCR instrument, Co-PAR enabled the codetection of multiplex proteins and RNAs, with the sensitivity of 102 EVs/µL (targeting CD63) and 1 EV/µL (targeting snRNA U6). We analyzed the coexpressions of three protein markers (CD63, GPC-1, HER2) and three RNA markers (snRNA U6, GPC-1 mRNA, miR-10b) on EVs from three pancreatic cell lines and 30 human plasma samples using Co-PAR. The diagnostic accuracy of the 6-biomarker combination reached 92.9%, which was at least 6.2% higher than that of 3-biomarker combinations and at least 13.5% higher than that of 6 single biomarkers. Co-PAR, as a multiparameter detection platform for EVs, has great potential in early disease diagnosis.
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Biomarcadores Tumorais , Detecção Precoce de Câncer , Vesículas Extracelulares , Neoplasias Pancreáticas , Humanos , Vesículas Extracelulares/química , Vesículas Extracelulares/metabolismo , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/metabolismo , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/análise , RNA/análise , Linhagem Celular TumoralRESUMO
Mass cytometry by time-of-flight (CyTOF), a high-dimensional single-cell analysis platform, detects up to 50 biomarkers at single-cell resolution. However, CyTOF analysis of biological samples with a minimal number of available cells or rare cell subsets remains a major technical challenge due to the extensive loss of cells during cell recovery, staining, and acquisition. Here, we introduce a platinum-chimeric carrier cell strategy for mass cytometry profiling of ultratrace cell samples. Cisplatin can rapidly enter broken plasma membranes of dead cells and form a chimeric interaction with cellular proteins, peptides, and amino acids. Thus, 198Pt-cisplatin is adopted to tag carrier cells in the pretreatment stage. We investigated 8 cell lines that are commonly accessible in laboratories for their potential as carrier cells to preserve rare target cells for CyTOF analysis. We designed a panel of 35 protein biomarkers to evaluate the comprehensive single-cell subtype classification capability with or without the carrier cell strategy. We further demonstrated the detection and analysis of as few as 1 × 104 immune cells using our method. The proposed method thus allows CyTOF analysis on precious clinical samples with less abundant cells.
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De novo design of peptides that bind specifically to functional proteins is beneficial for diagnostics and therapeutics. However, complex permutations and combinations of amino acids pose significant challenges to the rational design of peptides with desirable stability and affinity. Herein, we develop a computational-based evolution method, namely, peptidomimetics-driven recognition elements design (PepDRED), to derive hemoglobin-inspired peptidomimetics. PepDRED mimics the natural evolutionism pipeline to generate stable apovariant (AVs) structures for wild-type counterparts via automated point mutations and validates their efficiency through free binding energy analysis and per residue energy decomposition analysis. For application demonstration, we applied PepDRED to design de novo peptides to bind FhuA, a typical TonB-dependent transporter (TBDT). TBDTs are Gram-negative bacterial outer membrane proteins responsible for iron transport and vital for bacterial resistance. PepDRED generated a pool of AVs and proceeded to reach an optimized peptide, AV440, with a remarkable binding affinity of -21 kcal/mol. AV440 is â¼2.5-fold stronger than the existing FhuA inhibitor Microcin J25. Network energy analysis further unveils that incorporating methionine (M42) in the N-terminal region significantly enhances inter-residue contacts and binding affinity. PepDRED offers a prompt and efficient in silico approach to develop potent peptide candidates for target proteins.
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Proteínas de Escherichia coli , Peptidomiméticos , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Peptídeos/metabolismo , Ligação Proteica , Proteínas de Bactérias/químicaRESUMO
Pancreatic cancer, at unresectable advanced stages, presents poor prognoses, which could be prevented by early pancreatic cancer diagnosis methods. Recently, a promising early-stage pancreatic cancer biomarker, extracellular vesicles (EVs) related glypican-1 (GPC1) mRNA, is found to overexpress in pancreatic cancer cells. Current mRNA detection methods usually require expensive machinery, strict preservation environments, and time-consuming processes to guarantee detection sensitivity, specificity, and stability. Herein, we propose a novel two-step amplification method (CHAGE) via the target triggered Catalytic Hairpin Assembly strategy combined with Gold-Enhanced point-of-care-testing (POCT) technology for sensitive visual detection of pancreatic cancer biomarker. First, utilizing the catalyzed hairpin DNA circuit, low expression of the GPC1 mRNA was changed into amplification product 1 (AP1, a DNA duplex) as the next detection targets of the paper strips. Second, the AP1 was loaded onto a lateral flow assay and captured with the gold signal nanoparticles to visualize results. Finally, the detected results can be further enhanced by depositing gold to re-enlarge the sizes of gold nanoparticles in detection zones. As a result, the CHAGE methodology lowers the detection limit of mRNA to 100 fM and provides results within 2 h at 37 °C. Furthermore, we demonstrate the successful application in discriminating pancreatic cancer cells by analyzing EVs' GPC1 mRNA expression levels. Hence, the CHAGE methodology proposed here provides a rapid and convenient POCT platform for sensitive detection of mRNAs through unique probes designs (COVID, HPV, etc.).
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Detecção Precoce de Câncer/métodos , Neoplasias Pancreáticas/diagnóstico , RNA Mensageiro/isolamento & purificação , Biomarcadores Tumorais/genética , COVID-19 , Vesículas Extracelulares , Glipicanas/genética , Ouro , Humanos , Nanopartículas Metálicas , Neoplasias Pancreáticas/genética , Neoplasias PancreáticasRESUMO
Glucose, as an essential substance directly involved in metabolic processes, is closely related to the occurrence of various diseases such as glucose metabolism disorders and islet cell carcinoma. Therefore, it is crucial to develop sensitive, accurate, rapid, and cost effective methods for frequent and convenient detections of glucose. Microfluidic Paper-based Analytical Devices (µPADs) not only satisfying the above requirements but also occupying the advantages of portability and minimal sample consumption, have exhibited great potential in the field of glucose detection. This article reviews and summarizes the most recent improvements in glucose detection in two aspects of colorimetric and electrochemical µPADs. The progressive techniques for fabricating channels on µPADs are also emphasized in this article. With the growth of diabetes and other glucose indication diseases in the underdeveloped and developing countries, low-cost and reliably commercial µPADs for glucose detection will be in unprecedentedly demand.
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Glucose/análise , Dispositivos Lab-On-A-Chip , Técnicas Analíticas Microfluídicas/instrumentação , Papel , Colorimetria , EletroquímicaRESUMO
Antiviral vaccine is essential for preventing and controlling virus spreading, along with declining morbidity and mortality. A major challenge in effective vaccination lies in the ability to enhance both the humoral and cellular immune responses by adjuvants. Herein, self-assembled nanoparticles based on graphene oxide quantum dots with components of carnosine, resiquimod and Zn2+ ions, namely ZnGC-R, are designed as a new adjuvant for influenza vaccine. With its high capability for antigen-loading, ZnGC-R enhances antigen utilization, improves DC recruitment, and activates antigen-presenting cells. Single cell analysis of lymphocytes after intramuscular vaccination revealed that ZnGC-R generated multifaceted immune responses. ZnGC-R stimulated robust CD4+CCR7loPD-1hi Tfh and durable CD8+CD44hiCD62L- TEM immune responses, and simultaneously promoted the proliferation of CD26+ germinal center B cells. Besides, ZnGC-R elicited 2.53-fold higher hemagglutination-inhibiting antibody than commercial-licensed aluminum salt adjuvant. ZnGC-R based vaccine induced 342% stronger IgG antibody responses compared with vaccines with inactivated virus alone, leading to 100% in vivo protection efficacy against the H1N1 influenza virus challenge.
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Grafite , Vírus da Influenza A Subtipo H1N1 , Vacinas contra Influenza , Infecções por Orthomyxoviridae , Humanos , Adjuvantes Imunológicos/farmacologia , Imunidade Celular , Adjuvantes Farmacêuticos/farmacologia , Anticorpos Antivirais , Infecções por Orthomyxoviridae/prevenção & controleRESUMO
Patients with polycystic kidney disease (PKD) encounter a high risk of clear cell renal cell carcinoma (ccRCC), a malignant tumor with dysregulated lipid metabolism. SET domain-containing 2 (SETD2) has been identified as an important tumor suppressor and an immunosuppressor in ccRCC. However, the role of SETD2 in ccRCC generation in PKD remains largely unexplored. Herein, we perform metabolomics, lipidomics, transcriptomics and proteomics within SETD2 loss induced PKD-ccRCC transition mouse model. Our analyses show that SETD2 loss causes extensive metabolic reprogramming events that eventually results in enhanced sphingomyelin biosynthesis and tumorigenesis. Clinical ccRCC patient specimens further confirm the abnormal metabolic reprogramming and sphingomyelin accumulation. Tumor symptom caused by Setd2 knockout is relieved by myriocin, a selective inhibitor of serine-palmitoyl-transferase and sphingomyelin biosynthesis. Our results reveal that SETD2 deficiency promotes large-scale metabolic reprogramming and sphingomyelin biosynthesis during PKD-ccRCC transition. This study introduces high-quality multi-omics resources and uncovers a regulatory mechanism of SETD2 on lipid metabolism during tumorigenesis.
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Carcinoma de Células Renais , Neoplasias Renais , Animais , Camundongos , Humanos , Carcinoma de Células Renais/patologia , Esfingomielinas , Neoplasias Renais/patologia , Genes Supressores de Tumor , Transformação Celular Neoplásica/genética , Histona-Lisina N-MetiltransferaseRESUMO
Systemic Lupus Erythematosus (SLE) etiopathogenesis highlights the contributions of overproduction of CD4+ T cells and loss of immune tolerance. However, the involvement of CD8+ T cells in SLE pathology and disease progression remains unclear. Here, the comprehensive immune cell dysregulation in total 263 clinical peripheral blood samples composed of active SLE (aSLE), remission SLE (rSLE) and healthy controls (HCs) is investigated via mass cytometry, flow cytometry and single-cell RNA sequencing. This is observed that CD8+ CD27+ CXCR3- T cells are increased in rSLE compare to aSLE. Meanwhile, the effector function of CD8+ CD27+ CXCR3- T cells are overactive in aSLE compare to HCs and rSLE, and are positively associated with clinical SLE activity. In addition, the response of peripheral blood mononuclear cells (PBMCs) is monitored to interleukin-2 stimulation in aSLE and rSLE to construct dynamic network biomarker (DNB) model. It is demonstrated that DNB score-related parameters can faithfully predict the remission of aSLE and the flares of rSLE. The abundance and functional dysregulation of CD8+ CD27+ CXCR3- T cells can be potential biomarkers for SLE prognosis and concomitant diagnosis. The DNB score with accurate prediction to SLE disease progression can provide clinical treatment suggestions especially for drug dosage determination.
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Linfócitos T CD4-Positivos , Lúpus Eritematoso Sistêmico , Humanos , Linfócitos T CD8-Positivos , Leucócitos Mononucleares , Lúpus Eritematoso Sistêmico/diagnóstico , Biomarcadores , Progressão da Doença , Receptores CXCR3RESUMO
Lipopolysaccharide (LPS), also known as endotoxin, triggers a fatal septic shock; therefore, fast and accurate detection of LPS from a complex milieu is of primary importance. Several LPS affinity binders have been reported so far but few of them have proved their efficacy in developing electrochemical sensors capable of selectively detecting LPS from crude biological liquors. In this study, we identified 10 different single-stranded DNA aptamers showing specific affinity to LPS with dissociation constants (K(d)) in the nanomolar range using a NECEEM-based non-SELEX method. Based on the sequence and secondary structure analysis of the LPS binding aptamers, an aptamer exhibiting the highest affinity to LPS (i.e., B2) was selected to construct an impedance biosensor on a gold surface. The developed electrochemical aptasensor showed excellent sensitivity and specificity in the linear detection range from 0.01 to 1 ng/mL of LPS with significantly reduced detection time compared with the traditional Limulus amoebocyte lysate (LAL) assay.
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Aptâmeros de Nucleotídeos/química , Técnicas Eletroquímicas/métodos , Lipopolissacarídeos/análise , Aptâmeros de Nucleotídeos/genética , Sequência de Bases , Técnicas Biossensoriais , Impedância Elétrica , Eletrodos , Elétrons , Eletroforese Capilar , Biblioteca Gênica , Ouro/química , Cinética , Dados de Sequência Molecular , Conformação de Ácido NucleicoRESUMO
MicroRNAs (miRNAs), as the small, non-coding, evolutionary conserved, and post-transcriptional gene regulators of the genome, have been highly associated with various diseases such as cancers, viral infections, and cardiovascular diseases. Several techniques have been established to detect miRNAs, including northern blotting, real-time polymerase chain reaction (RT-PCR), and fluorescent microarray platform. However, it remains a significant challenge to develop sensitive, accurate, rapid, and cost-effective methods to detect miRNAs due to their short size, high similarity, and low abundance. The electrochemical biosensors exhibit tremendous potential in miRNA detection because they satisfy feature integration, portability, mass production, short response time, and minimal sample consumption. This article reviewed the working principles and signal amplification strategies of electrochemical DNA biosensors summarized the recent improvements. With the development of DNA nanotechnology, nanomaterials and biotechnology, electrochemical DNA biosensors of high sensitivity and specificity for microRNA detection will shortly be commercially accessible.
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The abuse application of glyphosate can result in a potential hazard for environment and human, however its ultrasensitive detection remains challenging. Herein, a Cu2+ modulated DNA-templated silver nanoclusters (DNA-AgNCs) sensor was constructed to sensitively determine glyphosate based on the turn-on fluorescence strategy. The fluorescence quenching of DNA-AgNCs occurred with the existence of Cu2+. Upon the presence of glyphosate, the functional groups on the surface of glyphosate could chelate with Cu2+, following the fluorescence recovery of DNA-AgNCs. Through the stoichiometric methods, we unveil that Cu2+-trigged fluorescence quenching mode is a combination of static and dynamic quenching with the static mode being predominant. In DNA-AgNCs/Cu2+ system, the carboxylate, amine, and phosphonate groups of glyphosate interact with Cu2+ through chelation, in which the carboxylate oxygen, the phosphonate oxygen atoms, and the monoprotonated secondary amine nitrogen atom and Cu2+ form chelate rings. This fluorescence sensor showed a desired linearity of glyphosate analysis under the optimum conditions, ranging from 15 to 100 µg/L with a low detection down to 5 µg/L. Moreover, the proposed sensor was successfully utilized to measure glyphosate in real samples, indicating a promising application in pesticide residues detection.
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Nanopartículas Metálicas , Prata , DNA/genética , Glicina/análogos & derivados , Humanos , GlifosatoRESUMO
The boosting exploitation of graphene oxide (GO) increases exposure risk to human beings. However, as primary defender in the first immune line, neutrophils' mechanism of defensive behavior toward GO remains unclear. Herein, we discovered that neutrophils recognize and defensively degrade GO in a lateral dimension dependent manner. The micrometer-sized GO (mGO) induces NETosis by releasing neutrophil extracellular traps (NETs), while nanometer-sized GO (nGO) elicits neutrophil degranulation. The two neutrophils' defensive behaviors are accompanied with generation of reactive oxygen species and activation of p-ERK and p-Akt kinases. However, mGO-induced NETosis is NADPH oxidase (NOX)-independent while nGO-triggered degranulation is NOX-dependent. Furthermore, myeloperoxidase (MPO) is determinant mediator despite distinct neutrophil phenotypes. Neutrophils release NETs comprising of MPO upon activated with mGO, while MPO is secreted via nGO-induced degranulation. Moreover, the binding energy between MPO and GO is calculated to be 69.8728 kJ mol-1 , indicating that electrostatic interactions mainly cause the spontaneous binding process. Meanwhile, the central enzymatic biodegradation occurs at oxygenic active sites and defects on GO. Mass spectrometry analysis deciphers the degradation products are biocompatible molecules like flavonoids and polyphenols. This study provides fundamental evidence and practical guidance for nanotechnology based on GO, including vaccine adjuvant, implantable devices, and energy storage.
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Armadilhas Extracelulares , Luta Romana , Grafite , Óxido de Magnésio/metabolismo , Neutrófilos , Espécies Reativas de Oxigênio/metabolismoRESUMO
Central nervous system diseases commonly occur with the destruction of the blood-brain barrier. As a primary cause of morbidity and mortality, stroke remains unpredictable and lacks cellular biomarkers that accurately quantify its occurrence and development. Here, we identify NeuN+/CD45-/DAPI+ phenotype nonblood cells in the peripheral blood of mice subjected to middle cerebral artery occlusion (MCAO) and stroke patients. Since NeuN is a specific marker of neural cells, we term these newly identified cells as circulating neural cells (CNCs). We find that the enumeration of CNCs in the blood is significantly associated with the severity of brain damage in MCAO mice (p < 0.05). Meanwhile, the number of CNCs is significantly higher in stroke patients than in negative subjects (p < 0.0001). These findings suggest that the amount of CNCs in circulation may serve as a clinical indicator for the real-time prognosis and progression monitor of the occurrence and development of ischemic stroke and other nervous system disease.
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Protein coating is a strategy for modifying and improving the surface functional properties of nanomaterials. However, the underlying mechanism behind protein coating formation, which is essential for its practical applications, remains largely unknown. Herein, we investigate the fundamental molecular mechanism of protein coating formation. Polydopamine nanospheres (PDANS) coated with bovine serum albumin (BSA) are examined in this study due to their wide biomedical potential. Our results demonstrate that BSAs can flexibly bind to PDANS and maintain their structural dynamicity. Our findings unveil that regular structure formation arises from BSAs lateral interactions via electrostatic forces. Notably, the protein coating modified PDANS surface enhances cell adhesion and proliferation as well as osteogenic differentiation. Such an enhancement is attributed to complementary surface properties provided by the dynamic PDANS-BSA complex and regular structure caused by BSA-BSA interactions in protein coating formation. This study provides a fundamental understanding of the molecular mechanism of protein coating formation, which facilitates the further development of functional protein-coated nanomaterials and guides the bioengineering decision making for biomedical applications, especially in bone tissue engineering.
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Nanosferas , Soroalbumina Bovina , Diferenciação Celular , Indóis , Osteogênese , PolímerosRESUMO
Typography-like templates for polydimethylsiloxane (PDMS) microfluidic chips using a fused deposition modeling (FDM) three-dimensional (3D) printer are presented. This rapid and fast proposed scheme did not require complicated photolithographic fabrication facilities and could deliver resolutions of ~100 µm. Polylactic acid (PLA) was adopted as the material to generate the 3D-printed units, which were then carefully assembled on a glass substrate using a heat-melt-curd strategy. This craft of bonding offers a cost-effective way to design and modify the templates of microfluidic channels, thus reducing the processing time of microfluidic chips. Finally, a flexible microfluidic chip to be employed for cell-based drug screening was developed based on the modularized 3D-printed templates. The lithography-free, typography-like, 3D-printed templates create a modularized fabrication process and promote the prevalence of integrated microfluidic systems with minimal requirements and improved efficiency.
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Dimetilpolisiloxanos , Ensaios de Seleção de Medicamentos Antitumorais/instrumentação , Técnicas Analíticas Microfluídicas/instrumentação , Poliésteres , Impressão Tridimensional , Células A549 , Cisplatino , HumanosRESUMO
DNA hydrogel garnered increasing attention in the sensing and medical field owing to its native biocompatibility and mechanical stability. While electrochemistry serves as a quantitative and sensitive detection technique, electrochemical DNA hydrogel biosensor is rarely reported. Here, for the first time, we report an electrochemical biosensor based on hybrid DNA hydrogel immobilized on indium tin oxide/polyethylene terephthalate (ITO/PET) electrode for the detection of lung cancer-specific microRNA, miR-21. The biosensor is capable of detecting miR-21 at a concentration as low as 5nM (1 pmol) and linear read-out from 10nM to 50µM. Ferrocene-tagged recognition probes were cross-linked with DNAs grafted on the polyacrylamide backbones to form the hybrid DNA hydrogel, which was further immobilized on 3-(trimethoxysilyl)propyl methacrylate (KH 570) treated ITO electrode. When the recognition probe was hybridized with the target miR-21, the hydrogel dissolved, producing a loss of ferrocene tags and a reduction in current, detected by Cyclic Voltammetry (CV) and Differential Pulse Voltammetry (DPV). The material characteristics of the biosensor were verified using contact angle meter and Energy Dispersive Spectrometer (EDS). This novel biosensor holds great promise in early sensitive clinical diagnosis for a variety of cancer-specific biomarkers due to the flexible sequence design of the recognition probe.
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Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/métodos , Neoplasias Pulmonares/diagnóstico , MicroRNAs/isolamento & purificação , Eletrodos , Humanos , Hidrogel de Polietilenoglicol-Dimetacrilato/química , Neoplasias Pulmonares/genética , Metacrilatos/química , MicroRNAs/genética , Hibridização de Ácido Nucleico/genética , Polietilenotereftalatos/química , Silanos/química , Compostos de Estanho/químicaRESUMO
Infections from parasitic nematodes (or roundworms) contribute to a significant disease burden and productivity losses for humans and livestock. The limited number of anthelmintics (or antinematode drugs) available today to treat these infections are rapidly losing their efficacy as multidrug resistance in parasites becomes a global health challenge. We propose an engineering approach to discover an anthelmintic drug combination that is more potent at killing wild-type Caenorhabditis elegans worms than four individual drugs. In the experiment, freely swimming single worms are enclosed in microfluidic drug environments to assess the centroid velocity and track curvature of worm movements. After analyzing the behavioral data in every iteration, the feedback system control (FSC) scheme is used to predict new drug combinations to test. Through a differential evolutionary search, the winning drug combination is reached that produces minimal centroid velocity and high track curvature, while requiring each drug in less than their EC50 concentrations. The FSC approach is model-less and does not need any information on the drug pharmacology, signaling pathways, or animal biology. Toward combating multidrug resistance, the method presented here is applicable to the discovery of new potent combinations of available anthelmintics on C. elegans, parasitic nematodes, and other small model organisms.
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Anti-Helmínticos/farmacologia , Caenorhabditis elegans/efeitos dos fármacos , Descoberta de Drogas/métodos , Testes de Sensibilidade Parasitária , Animais , Relação Dose-Resposta a Droga , Quimioterapia Combinada , Humanos , Testes de Sensibilidade Parasitária/métodosRESUMO
Endotoxin, present in the outer membrane of all gram-negative bacteria, can pose serious risks to human health, from irreversible shock to death. Therefore, it is essential to develop sensitive, accurate, and rapid methods for its detection. The rabbit pyrogen test is the first standard technique for endotoxin detection and, nowadays, has been replaced by the Limulus Amoebocyte Lysate test, which is the most popular detection technique for endotoxin. With in-depth understanding of endotoxin, biosensors based on endotoxin-sensing components are promising alternatives to pursue in developing low-cost, easy-operation, and fast-response endotoxin detection techniques. This article summarizes the recent advances of endotoxin detection methods with a particular emphasis on optical and electrochemical biosensors based on various sensing elements ranging from nature biomolecules to artificial materials. As the research and technological revolution continues, the highly integrated and miniaturized commercial devices for sensitively and reliably detecting endotoxin will provide a wide range of applications in people's daily life.
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Técnicas Biossensoriais , Endotoxinas/análise , Teste do Limulus , Animais , CoelhosRESUMO
Recent reports have indicated that aberrant expression of microRNAs is highly correlated with occurrence of lung cancer. Therefore, highly sensitive detection of lung cancer specific microRNAs provides an attractive approach in lung cancer early diagnostics. Herein, we designed 3D DNA origami structure that enables electrochemical detection of lung cancer related microRNAs. The 3D DNA origami structure is constituted of a ferrocene-tagged DNA of stem-loop structure combined with a thiolated tetrahedron DNA nanostructure at the bottom. The top portion hybridized with the lung cancer correlated microRNA, while the bottom portion was self-assembled on gold disk electrode surface, which was modified with gold nanoparticles (Au NPs) and blocked with mercaptoethanol (MCH). The preparation process and the performance of the proposed electrochemical genosensor were characterized by scanning electron microscopy (SEM), atomic force microscopy (AFM), cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Under the optimal conditions, the developed genosensor had a detection limit of 10 pM and a good linearity with microRNA concentration ranging from 100 pM to 1 µM, which showed a great potential in highly sensitive clinical cancer diagnosis application.
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Técnicas Biossensoriais/instrumentação , Ácidos Nucleicos Imobilizados/química , Neoplasias Pulmonares/genética , MicroRNAs/análise , Nanoestruturas/química , Hibridização de Ácido Nucleico , Técnicas Eletroquímicas/instrumentação , Desenho de Equipamento , Compostos Ferrosos/química , Ouro/química , Humanos , Ácidos Nucleicos Imobilizados/genética , Limite de Detecção , Pulmão/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/diagnóstico , Nanopartículas Metálicas/química , Metalocenos , MicroRNAs/genética , Nanoestruturas/ultraestruturaRESUMO
Dihydronicotinamide adenine dinucleotide (NADH) is a major biomarker correlated with lethal diseases such as cancers and bacterial infection. Herein, we report a graphene-DNA tetrahedron-gold nanoparticle modified gold disk electrode for highly sensitive NADH detection. By assembling the DNA tetrahedron/graphene composite film on the gold disk electrode surface which prior harnessed electrochemical deposition of gold nanoparticles to enhance the effective surface area, the oxidation potential of NADH was substantially decreased to 0.28V (vs. Ag/AgCl) and surface fouling effects were successfully eliminated. Furthermore, the lower detection limit of NADH by the presented platform was reduced down to 1fM, with an upper limit of 10pM. Both the regeneration and selectivity of composite film-modified electrode are investigated and proved to be robust. The novel sensor developed here could serve as a highly sensitive probe for NADH detection, which would further benefit the field of NADH related disease diagnostics.