RESUMEN
Developing non-passivating and fully integrated electrode arrays for point-of-care testing of carcinoembryonic antigen (CEA) is crucial, as the serum level of CEA is closely associated with colorectal cancer. Herein, we propose a simple, low-cost, and eco-friendly template-assisted filtration method for the scalable preparation of carbon nanotube-bridged Ti3C2Tx MXene (MX@CNT) electrode arrays with a conductive network. Furthermore, we fabricate a homogeneous electrochemical (HEC) sensor for CEA detection by integrating a magnetic-bead-based alkaline phosphatase-linked immunoassay (MB-aElisa), which enables the in-situ generation of the electroactive substance 1-naphthol (1-NP). Benefiting from the unique electrochemical characteristics of a MX@CNT electrode array, such as ultra-low background signal and superior electrocatalytic activity towards the hydrolyzed 1-NP, the MB-aElisa-based HEC sensor specifically measures CEA within a detection range spanning from 0.005 to 1.0 ng mL-1, achieving a detection limit of 1.6 pg mL-1. Subsequently, this biosensing prototype is successfully utilized for the detection of CEA in serum specimens obtained from colorectal cancer patients. More importantly, the integration of MB-aElisa with a MX@CNT electrode array not only marks a significant advancement but also enables the creation of a one-step homogeneous electrochemical immunosensing platform, serving as a paradigm for the highly sensitive and selective measurement of trace tumor markers in complex biological samples.
Asunto(s)
Biomarcadores de Tumor , Técnicas Biosensibles , Antígeno Carcinoembrionario , Técnicas Electroquímicas , Límite de Detección , Nanotubos de Carbono , Nanotubos de Carbono/química , Humanos , Técnicas Biosensibles/instrumentación , Antígeno Carcinoembrionario/sangre , Técnicas Electroquímicas/métodos , Biomarcadores de Tumor/sangre , Inmunoensayo/métodos , Inmunoensayo/instrumentación , Anticuerpos Inmovilizados/química , Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/sangre , ElectrodosRESUMEN
Early and rapid diagnostic of acute myocardial infarction (AMI) during its developing stage is crucial due to its high fatality rate. Heart-type fatty acid binding protein (h-FABP) is an ideal biomarker for the quantitative diagnosis of AMI, surpassing traditional markers such as myoglobin, creatine phosphokinase-MB, and troponin in terms of sensitivity, specificity, and prognostic value. To obtain diagnostic and prognostic information, a precise and fully quantitative measurement of h-FABP is essential, typically achieved through an immunosorbent assay like the enzyme-linked immunosorbent assay. Nevertheless, this method has several limitations, including extended detection time, complex assay procedures, the necessity for skilled technicians, and challenges in implementing automated detection. This research introduces a novel biosensor, utilizing aggregation-induced emission nanoparticles (AIENPs) and integrated with a digital microfluidic (DMF) workstation, designed for the sensitive, rapid, and automated detection of h-FABP in low-volume serum samples. AIENPs and magnetic beads in nanoscale were served as the capture particles and the fluorescent probe, which were linked covalently to anti-h-FABP antibodies respectively. The approach was based on a sandwich immunoassay and performed on a fully automated DMF workstation with assay time by 15 min. We demonstrated the determination of h-FABP in serum samples with detection limit of 0.14 ng/mL using this biosensor under optimal condition. Furthermore, excellent correlations (R2 = 0.9536, n = 50) were obtained between utilizing this biosensor and commercialized ELISA kits in clinical serum detecting. These results demonstrate that our flexible and reliable biosensor is suitable for direct integration into clinical diagnostics, and it is expected to be promising diagnostic tool for early detection and screening tests as well as prognosis evaluation for AMI patients.
Asunto(s)
Técnicas Biosensibles , Proteína 3 de Unión a Ácidos Grasos , Infarto del Miocardio , Nanopartículas , Técnicas Biosensibles/instrumentación , Humanos , Proteína 3 de Unión a Ácidos Grasos/sangre , Infarto del Miocardio/diagnóstico , Infarto del Miocardio/sangre , Nanopartículas/química , Límite de Detección , Biomarcadores/sangre , Proteínas de Unión a Ácidos Grasos/sangre , Proteínas de Unión a Ácidos Grasos/análisis , Inmunoensayo/métodos , Inmunoensayo/instrumentación , Microfluídica/métodos , Diseño de Equipo , Anticuerpos Inmovilizados/química , Ensayo de Inmunoadsorción EnzimáticaRESUMEN
A spatial-resolved and self-calibrated photoelectrochemical (PEC) biosensor has been fabricated by a multifunctional CeO2/CdS heterostructure, achieving portable and sensitive detection of carcinoembryonic antigen (CEA) using a homemade 3D printing device. The CeO2/CdS heterostructure with matched band structure is prepared to construct the dual-photoelectrodes to improve the PEC response of CeO2. In particular, as the photoactive nanomaterial, the CeO2 also plays the role of peroxidase mimetic nanozymes. Therefore, the catalytic performance of CeO2 with different morphologies (e.g., nano-cubes, nano-rods and nano-octahedra) have been studied, and CeO2 nano-cubes (c-CeO2) achieve the optimal catalytic activity. Upon introducing CEA, the sandwich-type immunocomplex is formed in the microplate using GOx-AuNPs-labeled second antibody as detection antibody. As a result, H2O2 can be produced from the catalytic oxidization of glucose substrate by GOx, which is further catalyzed by CeO2 to form â¢OH, thus in situ etching CdS and decreasing the photocurrents. The self-calibration is achieved by the dual-channel photoelectrodes on the homemade 3D printing device to obtain the photocurrents ratio, thus effectively normalizing the fluctuations of external factors to enhance the accuracy. This integrated biosensor with a detection limit as low as 0.057 ng mL-1 provides a promising way for ultrasensitive immunoassay in clinic application in complex environments.
Asunto(s)
Técnicas Biosensibles , Compuestos de Cadmio , Antígeno Carcinoembrionario , Cerio , Técnicas Electroquímicas , Impresión Tridimensional , Sulfuros , Técnicas Biosensibles/instrumentación , Cerio/química , Inmunoensayo/instrumentación , Inmunoensayo/métodos , Antígeno Carcinoembrionario/sangre , Compuestos de Cadmio/química , Sulfuros/química , Humanos , Límite de Detección , Oro/química , Anticuerpos Inmovilizados/química , Nanopartículas del Metal/químicaRESUMEN
BACKGROUND: Cardiac troponin I (CTnI) is demonstrated as one of the most promising disease biomarkers for early diagnosing acute myocardial infarction (AMI). To date, electrochemical immunosensors have been extensively studied in the field of cTnI determination. But highly accurate and sensitive cTnI detection by this method is still a challenge due to non-specific adsorption on electrode interfaces in complex human serum. As a result, it is necessary to develop an antifouling electrochemical immunosensor with high sensitivity for the detection of cTnI. RESULTS: In this work, an antifouling electrochemical immunosensor was constructed based on vertically-aligned peptide layer consisting of Au nanoparticles (AuNPs) and amphiphilic CEAK16 peptide (CEAK16@AuNPs) for sensitive and accurate detection of cTnI in human serum. The vertically-aligned CEAK16@AuNPs interface provided a stable hydration layer originated from attraction of water molecules by amino acids on the hydrophilic side of the CEAK16, which effectively reduced non-specific adsorption and enhanced electron transfer rate. The cTnI immunosensor possessed great analytical performance with a wide range from 1 fg mL-1 to 1 µg mL-1 and a low detection limit of 0.28 fg mL-1 (S/N = 3). Additionally, the proposed CEAK16@AuNPs sensing interface showed excellent long-term antifouling performance and electrochemical activity that preserved 80 % of the initial signal after 20-days exposure in human serum samples. Consequently, the cTnI immunosensor displayed excellent detection accuracy compared to clinical methods and owned good selectivity, stability and reproducibility. SIGNIFICANCE: The development of this strategy provides a versatile tool for accurate quantitative cTnI analysis in real human serum, thus helping to achieve early AMI diagnosis effectively and holding the promising potentials for other immunosensor in disease diagnosis.
Asunto(s)
Técnicas Electroquímicas , Oro , Nanopartículas del Metal , Troponina I , Humanos , Troponina I/sangre , Oro/química , Nanopartículas del Metal/química , Límite de Detección , Técnicas Biosensibles , Péptidos/química , Inmunoensayo/métodos , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/química , ElectrodosRESUMEN
Detecting chronic autoimmune disorders (ADs) early reduces the risk of morbidity, disability, and mortality and offers the possibility of significant therapeutic action in a timely manner. Developing low-cost, reliable, and sensitive sensors for ADs can ensure the efficient utilization of healthcare resources at earlier stages. Here, we report on the development of an electrochemical biosensor for sensing CXCL10, a chemokine protein that serves as a biomarker for autoimmune diseases. A self-assembly strategy is used for the immobilization of biorecognition elements on a plastic chip electrode (PCE). A homemade PCE offers a versatile and cost-effective scaffold for sensing applications. Gold nanoparticles were electrochemically deposited on the electrode via the reduction of gold ions on the PCE galvanostatically. The CXCL10 antibody and recognition elements were immobilized on the gold-deposited PCE. The attachment of recognition molecules was confirmed by energy-dispersive scanning electron microscopy, atomic force microscopy, infrared spectroscopy, and electrochemical techniques. Electrochemical impedance spectroscopy (EIS) was used for the detection of CXCL10 within a concentration range spanning from pico- to micro-molar levels. The sensor exhibited remarkable linearity in both buffer and plasma solutions, with a limit of detection (LOD) of up to 0.72 pg mL-1.
Asunto(s)
Enfermedades Autoinmunes , Técnicas Biosensibles , Quimiocina CXCL10 , Técnicas Electroquímicas , Electrodos , Oro , Nanopartículas del Metal , Técnicas Biosensibles/métodos , Técnicas Biosensibles/instrumentación , Humanos , Enfermedades Autoinmunes/diagnóstico , Enfermedades Autoinmunes/sangre , Enfermedades Autoinmunes/inmunología , Oro/química , Nanopartículas del Metal/química , Quimiocina CXCL10/análisis , Quimiocina CXCL10/sangre , Quimiocina CXCL10/inmunología , Técnicas Electroquímicas/métodos , Técnicas Electroquímicas/instrumentación , Diagnóstico Precoz , Plásticos/química , Espectroscopía Dieléctrica/métodos , Límite de Detección , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/químicaRESUMEN
Lung cancer is the main cancer that endangers human life worldwide, with the highest mortality rate. The detection of lung tumor markers is of great significance for the early diagnosis and subsequent treatment of lung cancer. In this study, a vertical graphene field effect transistor (VGFET) immunosensor based on graphene/C60 heterojunction was created to offer quantitative detections for the lung tumor markers carcinoembryonic antigen (CEA), cytokeratin 19 fragment (Cyfra21-1), and neuron-specific enolase (NSE). The experimental results showed that the sensitive range for standard antigen is between 1 pg/ml to 100 ng/ml, with a limit of detection (LOD) of 5.6 amol/ml for CEA, 33.3 amol/ml for Cyfra 21-1 and 12.8 amol/ml for NSE (1 pg/ml for all). The detection accuracy for these tumor markers was compared with the clinically used method for clinical patients on serum samples. Results are highly consistent with clinically used immunoassay in its efficient diagnosis concentration range. Subsequently, the mesoporous silica nanospheres (MSNs) with an average size of 90 nm were surface modified with glutaraldehyde, and a second antibody was assembled on MSNs, which fixes nanospheres on the antigen and amplified the field effect. The LODs for three markers are 100 fg/ml (0.56 amol/ml for CEA) under optimal circumstances of detection. This result indicates that specific binding to MSNs enhances local field effects and can achieve higher sensing efficiency for tumor marker detection at extremely low concentrations, providing effective assistance for the early diagnosis of lung cancer.
Asunto(s)
Antígenos de Neoplasias , Biomarcadores de Tumor , Técnicas Biosensibles , Antígeno Carcinoembrionario , Grafito , Queratina-19 , Neoplasias Pulmonares , Fosfopiruvato Hidratasa , Grafito/química , Humanos , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/análisis , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/sangre , Queratina-19/sangre , Antígeno Carcinoembrionario/sangre , Técnicas Biosensibles/métodos , Fosfopiruvato Hidratasa/sangre , Inmunoensayo/métodos , Antígenos de Neoplasias/sangre , Antígenos de Neoplasias/análisis , Límite de Detección , Dióxido de Silicio/química , Transistores Electrónicos , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/química , Nanosferas/químicaRESUMEN
Nuclear matrix protein 22 (NMP22) is one of the most important tumor markers of bladder cancer and is significantly elevated in the urine of bladder cancer patients. Therefore, in this work, a highly sensitive ratiometric electrochemical immunosensor was constructed to detect NMP22 based on ZIF-8@MWCNTs@Chit@Fc@AuNPs composites. ZIF-8 had a large surface area and good adsorption ability. Multi-Walled Carbon Nanotubes (MWCNTs) can optimize the electrical conductivity of ZIF-8, so that the electrode surface of ferrocene (Fc) obtains a stable and strong electrochemical signal. In addition, AuPt-MB provided another strong detection signal methylene blue (MB) while immobilizing the secondary antibody (Ab2) through Au-N and Pt-N bonds. A ratiometric electrochemical sensor was formed based on ZIF-8@MWCNTs@Chit@Fc@AuNPs and AuPt-MB, which showed a great linear connection between IMB/IFc and the logarithmic concentration of NMP22 with a detection limit of 3.33 fg mL-1 (S/N = 3) under optimized specifications in the concentration interval of 0.01 pg mL-1 to 1000 ng mL-1. In addition, the ratiometric immunosensor showed good selectivity and stability.
Asunto(s)
Técnicas Biosensibles , Técnicas Electroquímicas , Oro , Nanopartículas del Metal , Nanotubos de Carbono , Proteínas Nucleares , Nanotubos de Carbono/química , Oro/química , Técnicas Electroquímicas/métodos , Humanos , Nanopartículas del Metal/química , Inmunoensayo/métodos , Técnicas Biosensibles/métodos , Proteínas Nucleares/orina , Proteínas Nucleares/inmunología , Proteínas Nucleares/análisis , Límite de Detección , Platino (Metal)/química , Zeolitas/química , Azul de Metileno/química , Compuestos Ferrosos/química , Anticuerpos Inmovilizados/química , Anticuerpos Inmovilizados/inmunología , Metalocenos/química , Biomarcadores de Tumor/orina , Biomarcadores de Tumor/inmunologíaRESUMEN
Human epididymis protein 4 (HE4), a diagnostic biomarker of ovarian cancer, is crucial for monitoring the early stage of the disease. Hence, it is highly important to develop simple, inexpensive, and user-friendly biosensors for sensitive and quantitative HE4 assays. Herein, a new sandwich-type electrochemical immunosensor based on Prussian blue (PB) as a signal indicator and functionalized metal-organic framework nanocompositesas efficient signal amplifiers was fabricated for quantitative analysis of HE4. In principle, ketjen black (KB) and AuNPs modified on TiMOF (TiMOF-KB@AuNPs) could accelerate electron transfer on the electrode surface and act as a matrix for the immobilization of antibodies via cross-linking to improve the determination sensitivity. The PB that covalently binds to labeled antibodies endows the biosensors with intense electrochemical signals. Furthermore, the concentration of HE4 could be indirectly detected by monitoring the electroactivity of PB. Benefiting from the high signal amplification ability of the PB and MOF nanocomposites, this strategy displayed a wide linear range (0.1-80 ng mL-1) and a lower detection limit (0.02 ng mL-1). Hence, this study demonstrated great promise for application in clinical ovarian cancer diagnosis and treatment, and provided a new platform for detecting other cancer biomarkers.
Asunto(s)
Técnicas Biosensibles , Técnicas Electroquímicas , Ferrocianuros , Oro , Límite de Detección , Estructuras Metalorgánicas , Neoplasias Ováricas , Proteína 2 de Dominio del Núcleo de Cuatro Disulfuros WAP , Técnicas Biosensibles/métodos , Humanos , Estructuras Metalorgánicas/química , Proteína 2 de Dominio del Núcleo de Cuatro Disulfuros WAP/análisis , Ferrocianuros/química , Técnicas Electroquímicas/métodos , Neoplasias Ováricas/diagnóstico , Neoplasias Ováricas/sangre , Femenino , Oro/química , Nanopartículas del Metal/química , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/análisis , Inmunoensayo/métodos , Anticuerpos Inmovilizados/química , Nanocompuestos/químicaRESUMEN
Biosensors based on immobilized antibodies require molecular strategies that (i) couple the antibodies in a stable fashion while maintaining the conformation and functionality, (ii) give outward orientation of the paratope regions of the antibodies for good accessibility to analyte molecules in the biofluid, and (iii) surround the antibodies by antibiofouling molecules. Here, we demonstrate a method to achieve oriented coupling of antibodies to an antifouling poly(l-lysine)-grafted-poly(ethylene glycol) (PLL-g-PEG) substrate, using glycan remodeling to create antibody-DNA conjugates. The coupling, orientation, and functionality of the antibodies were studied using two analysis methods with single-molecule resolution, namely single-molecule localization microscopy and continuous biosensing by particle motion. The biosensing functionality of the glycan-remodeled antibodies was demonstrated in a sandwich immunosensor for procalcitonin. The results show that glycan-remodeled antibodies enable oriented immobilization and biosensing functionality with low nonspecific binding on antifouling polymer substrates.
Asunto(s)
Anticuerpos Inmovilizados , Técnicas Biosensibles , Polisacáridos , Técnicas Biosensibles/métodos , Polisacáridos/química , Polisacáridos/inmunología , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/química , Polietilenglicoles/química , Incrustaciones Biológicas/prevención & control , Polilisina/química , Anticuerpos/inmunología , Anticuerpos/química , Humanos , Polímeros/químicaRESUMEN
An electrochemical biosensor has been developed for detection of Escherichia coli O157 by integrating lateral flow with screen-printed electrodes. The screen-printed electrodes were attached under the lateral flow detection line, and organic-inorganic nanoflowers prepared from E. coli O157-specific antibodies as an organic component were attached to the lateral flow detection line. In the presence of E. coli O157, an organic-inorganic nanoflower-E. coli O157-antimicrobial peptide-labelled ferrocene sandwich structure is formed on the lateral flow detection line. Differential pulse voltammetry is applied using a smartphone-based device to monitor ferrocene on the detection line. The resulting electrochemical biosensor could specifically detect E. coli O157 with a limit of detection of 25 colony-forming units mL-1. Through substitution of antibodies of organic components in organic-inorganic nanoflowers, biosensors have great potential for the detection of other pathogens in biomedical research and clinical diagnosis.
Asunto(s)
Técnicas Biosensibles , Técnicas Electroquímicas , Escherichia coli O157 , Escherichia coli O157/aislamiento & purificación , Escherichia coli O157/inmunología , Técnicas Biosensibles/métodos , Inmunoensayo/métodos , Inmunoensayo/instrumentación , Técnicas Electroquímicas/métodos , Técnicas Electroquímicas/instrumentación , Límite de Detección , Nanoestructuras/química , Electrodos , Compuestos Ferrosos/química , Anticuerpos Inmovilizados/inmunología , Metalocenos/química , Anticuerpos Antibacterianos/química , Anticuerpos Antibacterianos/inmunología , Péptidos Antimicrobianos/químicaRESUMEN
An intelligent colorimetric sensing platform integrated with in situ immunomagnetic separation function was developed for ultrasensitive detection of Escherichia coli O157: H7 (E. coli O157: H7) in food. Captured antibody modified magnetic nanoparticles (cMNPs) and detection antibody/horseradish peroxidase (HRP) co-functionalized AuNPs (dHAuNPs) were firstly synthesized for targeted enrichment and colorimetric assay of E. coli O157: H7, in which remarkable signal amplification was realized by loading large amounts of HRP on the surface of AuNPs. Coupling with the optical collimation attachments and embedded magnetic separation module, a highly integrated optical device was constructed, by which in situ magnetic separation and high-quality imaging of 96-well microplates containing E. coli O157: H7 was achieved with a smartphone. The concentration of E. coli O157: H7 could be achieved in one-step by performing digital image colorimetric analysis of the obtained image with a custom-designed app. This biosensor possesses high sensitivity (1.63 CFU/mL), short detecting time (3 h), and good anti-interference performance even in real-sample testing. Overall, the developed method is expected to be a novel field detection platform for foodborne pathogens in water and food as well as for the diagnosis of infections due to its portability, ease of operation, and high feasibility.
Asunto(s)
Técnicas Biosensibles , Colorimetría , Escherichia coli O157 , Microbiología de Alimentos , Oro , Peroxidasa de Rábano Silvestre , Separación Inmunomagnética , Nanopartículas del Metal , Escherichia coli O157/aislamiento & purificación , Colorimetría/métodos , Oro/química , Peroxidasa de Rábano Silvestre/química , Separación Inmunomagnética/métodos , Técnicas Biosensibles/métodos , Nanopartículas del Metal/química , Contaminación de Alimentos/análisis , Límite de Detección , Teléfono Inteligente , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/química , Nanopartículas de Magnetita/químicaRESUMEN
The rampant hepatitis B virus (HBV) seriously endangers human health, and hepatitis B surface antigen (HBsAg) is its early diagnostic marker. Therefore, it is crucial to construct a fast and highly sensitive HBsAg detection method. Based on high-efficiency magnetic separation technology and fluorescent composite material labelling technology, an accurate, fast and sensitive fluorescent immunosensing system for HBsAg detection was developed. Immunomagnetic beads constructed from carboxyl-functionalized Fe3O4 nanoparticles (Fe3O4-COOH) with excellent magnetic response performance were used as efficient capture carriers for HBsAg. Immunofluorescence composite microspheres constructed based on ultra-stable polystyrene-coated CsPbBr3 perovskite nanocrystals (CPB@PSAA) with high hydrophilic properties, were excellent fluorescent markers for HBsAg. Using this sensitive sandwich fluorescence sensing system a good linear relationship within the range of 0.2-15 ng/mL was established between HBsAg concentration and fluorescence intensity with a limit of detection (LOD) of 0.05 ng/mL. The system obtained satisfactory results when tested on real human serum samples. The magnetic-assisted fluorescence immune-sandwich sensor system has broad application prospects in biomedicine such as rapid and early diagnosis and effective prevention of infectious diseases.
Asunto(s)
Compuestos de Calcio , Antígenos de Superficie de la Hepatitis B , Interacciones Hidrofóbicas e Hidrofílicas , Límite de Detección , Óxidos , Titanio , Antígenos de Superficie de la Hepatitis B/sangre , Antígenos de Superficie de la Hepatitis B/inmunología , Antígenos de Superficie de la Hepatitis B/análisis , Humanos , Óxidos/química , Titanio/química , Compuestos de Calcio/química , Colorantes Fluorescentes/química , Nanopartículas de Magnetita/química , Microesferas , Anticuerpos Inmovilizados/inmunología , Virus de la Hepatitis B/inmunología , Virus de la Hepatitis B/aislamiento & purificación , Inmunoensayo/métodosRESUMEN
A novel biofuel cell (BFC)-based self-powered electrochemical immunosensing platform was developed by integrating the target-induced biofuel release and biogate immunoassay for ultrasensitive 17ß-estradiol (E2) detection. The carbon nanocages/gold nanoparticle composite was employed in the BFCs device as the electrode material, through which bilirubin oxidase and glucose oxidase were wired to form the biocathode and bioanode, respectively. Positively charged mesoporous silica nanoparticles (PMSN) were encapsulated with glucose molecules as biofuel and subsequently coated by the negatively charged AuNPs-labelled anti-E2 antibody (AuNPs-Ab) serving as a biogate. The biogate could be opened efficiently and the trapped glucose released once the target E2 was recognized and captured by AuNPs-Ab due to the decreased adhesion between the antigen-antibody complex and PMSN. Then, glucose oxidase oxidized the glucose to produce a large number of electrons, resulting in significantly increased open-circuit voltage (EOCV). Promisingly, the proposed BFC-based self-powered immunosensor demonstrated exceptional sensitivity for the detection of E2 in the concentration range from 1.0 pg mL-1 to 10.0 ng mL -1, with a detection limit of 0.32 pg mL-1 (S/N = 3). Furthermore, the prepared BFC-based self-powered homogeneous immunosensor showed significant potential for implementation as a viable prototype for a mobile and an on-site bioassay system in food and environmental safety applications.
Asunto(s)
Fuentes de Energía Bioeléctrica , Técnicas Biosensibles , Estradiol , Glucosa Oxidasa , Oro , Límite de Detección , Nanopartículas del Metal , Inmunoensayo/métodos , Estradiol/química , Estradiol/análisis , Oro/química , Glucosa Oxidasa/química , Técnicas Biosensibles/métodos , Nanopartículas del Metal/química , Técnicas Electroquímicas/métodos , Técnicas Electroquímicas/instrumentación , Humanos , Electrodos , Glucosa/análisis , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/química , Anticuerpos Inmovilizados/inmunología , Dióxido de Silicio/química , Enzimas Inmovilizadas/químicaRESUMEN
An electrochemical immunoassay system was developed to detect CA-125 using a glassy carbon electrode (GCE) modified with MXene, graphene quantum dots (GQDs), and gold nanoparticles (AuNPs). The combined MXene-GQD/AuNPs modification displayed advantageous electrochemical properties due to the synergistic effects of MXene, GQDs, and AuNPs. The MXene-GQD composite in the modified layer provided strong mechanical properties and a large specific surface area. Furthermore, the presence of AuNPs significantly improved conductivity and facilitated the binding of anti-CA-125 on the modified GCE, thereby enhancing sensitivity. Various analytical techniques such as FE-SEM and EDS were utilized to investigate the structural and morphological characteristics as well as the elemental composition. The performance of the developed immunosensor was assessed using electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV), square wave voltammetry (SWV), and differential pulse voltammetry (DPV). Under optimized conditions in a working potential range of -0.2 to 0.6 V (vs. Ag/AgCl), the sensitivity, linear range (LR), limit of detection (LOD), and correlation coefficient (R2) were determined to be 315.250 µA pU.mL-1/cm2, 0.1 to 1 nU/mL, 0.075 nU/mL, and 0.9855, respectively. The detection of CA-125 in real samples was investigated using the developed immunoassay platform, demonstrating satisfactory results including excellent selectivity and reproducibility.
Asunto(s)
Antígeno Ca-125 , Técnicas Electroquímicas , Oro , Grafito , Límite de Detección , Nanopartículas del Metal , Neoplasias Ováricas , Puntos Cuánticos , Antígeno Ca-125/sangre , Antígeno Ca-125/análisis , Oro/química , Nanopartículas del Metal/química , Humanos , Neoplasias Ováricas/sangre , Técnicas Electroquímicas/métodos , Técnicas Electroquímicas/instrumentación , Inmunoensayo/métodos , Femenino , Puntos Cuánticos/química , Grafito/química , Anticuerpos Inmovilizados/inmunología , Técnicas Biosensibles/métodos , Electrodos , Proteínas de la MembranaRESUMEN
BACKGROUND: Okadaic acid (OA), as a diarrhetic shellfish poisoning, can increase the risk of acute carcinogenic or teratogenic effects for the ingestion of OA contaminated shellfish. At present, much effort has been made to graft immunoassay onto a paper substrate to make paper-based sensors for rapid and simple detection of shellfish toxin. However, the complicated washing steps and low protein fixation efficiency on the paper substrate need to be further addressed. RESULTS: A novel paper-tip immunosensor for detecting OA was developed combined with smartphone and naked eye readout. The trapezoid paper tip was consisted of quantitative and qualitative detection zones. To improve the OA antigen immobilization efficiency on the paper substrate, graphene oxide (GO)-assisted protein immobilization method was introduced. Meanwhile, Au nanoparticles composite probe combined with the lateral flow washing was developed to simplify the washing step. The OA antigen-immobilized zone, as the detection zone â , was used for quantitative assay by smartphone imaging. The paper-tip front, as the detection zone â ¡, which could qualitatively differentiate OA pollution level within 45 min using the naked eye. The competitive immunoassay on the paper tip exhibited a wide linear range for detecting OA (0.02-50 ngâmL-1) with low detection limit of 0.02 ngâmL-1. The recovery of OA in spiked shellfish samples was in the range of 90.3 %-113.%. SIGNIFICANCE: These results demonstrated that the proposed paper-tip immunosensor could provide a simple, low-cost and high-sensitivity test for OA detection without the need for additional large-scale equipment or expertise. We anticipate that this paper-tip immunosensor will be a flexible and versatile tool for on-site detecting the pollution of marine products.
Asunto(s)
Técnicas Biosensibles , Oro , Grafito , Ácido Ocadaico , Papel , Teléfono Inteligente , Grafito/química , Ácido Ocadaico/análisis , Inmunoensayo/métodos , Oro/química , Nanopartículas del Metal/química , Proteínas Inmovilizadas/química , Límite de Detección , Animales , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/químicaRESUMEN
In cryo-electron microscopy (cryo-EM), sample preparation poses a critical bottleneck, particularly for rare or fragile macromolecular assemblies and those suffering from denaturation and particle orientation distribution issues related to air-water interface. In this study, we develop and characterize an immobilized antibody-based affinity grid (IAAG) strategy based on the high-affinity PA tag/NZ-1 antibody epitope tag system. We employ Pyr-NHS as a linker to immobilize NZ-1 Fab on the graphene oxide or carbon-covered grid surface. Our results demonstrate that the IAAG grid effectively enriches PA-tagged target proteins and overcomes preferred orientation issues. Furthermore, we demonstrate the utility of our IAAG strategy for on-grid purification of low-abundance target complexes from cell lysates, enabling atomic resolution cryo-EM. This approach greatly streamlines the purification process, reduces the need for large quantities of biological samples, and addresses common challenges encountered in cryo-EM sample preparation. Collectively, our IAAG strategy provides an efficient and robust means for combined sample purification and vitrification, feasible for high-resolution cryo-EM. This approach holds potential for broader applicability in both cryo-EM and cryo-electron tomography (cryo-ET).
Asunto(s)
Anticuerpos Inmovilizados , Microscopía por Crioelectrón , Microscopía por Crioelectrón/métodos , Anticuerpos Inmovilizados/química , Anticuerpos Inmovilizados/inmunología , Grafito/química , HumanosRESUMEN
The ELISA is the most worldwide method for immunoassay. However, the ELISA is losing ground due to low reproducibility of manual experimental processes in both R&D and IVD areas. An automated platform is a good solution, but there are still limitations owning to extremely high cost and requiring large space to set up especially for a small size laboratory. Here, we present a novel all-in-one platform called "VEUS" settable on the laboratory table that offers comprehensive automation of the entire multiplex immunoassay process by exploiting antibody conjugated magnetic particles, quality control and then immunoanalytical reaction, thereby enhancing detection sensitivity and high reproducibility. As a proof of concept, the system exhibits a sensitive LOD of 0.6 and 3.1 pg mL-1 within 1 h run, comparable precision that of molecular diagnostic systems based on PCR method, enabling rapid multiplex diagnosis of Influenza A, Influenza B, and COVID-19 viruses with similar symptoms. Through automation by the all-in-one system, it can be used by novice users, something innovative for immunoassays, relying heavily on user experience. Furthermore, it can contribute to streamline entire immunoassay processes of diverse biomarkers with high reproducibility and convenience in laboratories.
Asunto(s)
SARS-CoV-2 , Humanos , Inmunoensayo/métodos , SARS-CoV-2/inmunología , SARS-CoV-2/aislamiento & purificación , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/química , Reproducibilidad de los Resultados , COVID-19/diagnóstico , COVID-19/virología , Ensayo de Inmunoadsorción Enzimática/métodos , Virus de la Influenza A/inmunología , Virus de la Influenza B/inmunología , Automatización de Laboratorios/métodos , Límite de DetecciónRESUMEN
Biosensors have become promising alternatives to the conventional methods in early identification of diseases. However, translation of biosensors from lab to commercial products have challenges such as complex sensor fabrications and complicated detection, and inadequate sensitivity and selectivity. Here, we introduce simple and low-cost fabricated conductometric sensors based on high resistivity silicon wafers (HR-Si) which can be adopted to functionalise with both natural and synthetic antibodies in detecting five biomarkers including interleukin-6, C reactive protein, cardiac troponin I, brain natriuretic peptide, and N terminal-probrain natriuretic peptide. All five biomarkers show selective and rapid (10 min sample incubation and <1 min of reading time) detection in both media of phosphate buffer saline and saliva with the detection limits lower than that of reported healthy levels in saliva. This work highlights the versatility of HR-Si sensors in functionalisation of both natural and synthetic antibodies in sensitive and selective biomarker detection. As these miniaturised conductometric biosensors can be easily modified with on-demand biomaterials to detect corresponding target biomarkers, they enable a new category of compact point-of-care medical devices.
Asunto(s)
Biomarcadores , Técnicas Biosensibles , Péptido Natriurético Encefálico , Saliva , Troponina I , Técnicas Biosensibles/instrumentación , Técnicas Biosensibles/métodos , Biomarcadores/análisis , Saliva/química , Humanos , Troponina I/análisis , Péptido Natriurético Encefálico/análisis , Proteína C-Reactiva/análisis , Límite de Detección , Interleucina-6/análisis , Diseño de Equipo , Silicio/química , Fragmentos de Péptidos/análisis , Anticuerpos Inmovilizados/química , Inflamación/diagnósticoRESUMEN
Aflatoxin B1 (AFB1), classified as a class I carcinogen, is a widespread mycotoxin that poses a serious threat to public health and economic development, and the food safety problems caused by AFB1 have aroused worldwide concern. The development of accurate and sensitive methods for the detection of AFB1 is significant for food safety monitoring. In this work, a sandwich-type photoelectrochemical (PEC) biosensor for AFB1 detection was constructed on the basis of an aptamer-antibody structure. A good photocurrent response was obtained due to the sensitization of In2S3 by Ru(bpy)32+. In addition, this sandwich-type sensor constructed by modification with the antibody, target detector, and aptamer layer by layer attenuated the migration hindering effect of photogenerated carriers caused by the double antibody structure. The aptamer and antibody synergistically recognized and captured the target analyte, resulting in more reliable PEC response signals. CdSe@CdS QDs-Apt were modified as a signal-off probe onto the sensor platform to quantitatively detect AFB1 with a "signal-off" response, which enhanced the sensitivity of the sensor. The PEC biosensor showed a linear response range from 10-12 to 10-6 g mL-1 with a detection limit of 0.023 pg mL-1, providing a feasible approach for the quantitative detection of AFB1 in food samples.
Asunto(s)
Aflatoxina B1 , Aptámeros de Nucleótidos , Técnicas Biosensibles , Técnicas Electroquímicas , Límite de Detección , Aflatoxina B1/análisis , Aflatoxina B1/inmunología , Técnicas Biosensibles/métodos , Aptámeros de Nucleótidos/química , Técnicas Electroquímicas/métodos , Técnicas Electroquímicas/instrumentación , Puntos Cuánticos/química , Contaminación de Alimentos/análisis , Compuestos de Cadmio/química , Anticuerpos Inmovilizados/inmunología , Anticuerpos Inmovilizados/química , Procesos Fotoquímicos , Sulfuros/química , Compuestos de Selenio/química , Compuestos OrganometálicosRESUMEN
Nanosized sodium bismuth perovskite titanate (NBT) was synthesized and first used as the electrochemical immune sensing platform for the sensitive detection of carcinoembryonic antigen (CEA). Gold nanoparticles (Au NPs) grew on the surface of NBT through forming Au-N bond to obtain Au@NBT, and a label-free electrochemical immunosensor was proposed using Au@NBT as an immunosensing recognizer towards CEA. The well-ordered crystal structure of NBT was not changed at all after the modification of Au NPs outside, but significantly improved the conductivity, catalytic activity, and biocompatibility of the Au@NBT-modified electrode. The unique cubic crystal nanostructure of NBT offered a large active area for both Au NP modification and the subsequent immobilization of biomolecules over the electrode surface, triggering the effective generation of promising properties of the proposed Au@NBT-based electrochemical immunosensor. As expected, favorable detection performances were achieved using this immunosensor towards CEA detection, where a good linear relationship between the current response and CEA concentration was obtained in the concentration range 10 fg mL-1 to 100 ng mL-1 with a low detection limit (LOD) of 13.17 fg mL-1. Also, the significantly enhanced selectivity, and stability guaranteed the promising electrochemical properties of this immunosensor. Furthermore, the analysis of real serum samples verified the high feasibility of this new method in clinical CEA detection. This work opens a new window for the application of nanoperovskite in the early detection of CEA.