RESUMEN
Tetracyclines are currently the most commonly used class of antibiotics, and their residue issue significantly impacts public health safety. In this study, a surface modification of perovskite with cetyltrimethylammonium bromide led to the generation of stable electrochemiluminescence (ECL) emitters in aqueous systems and improved the biocompatibility of perovskite. A perovskite quantum dot-based ECL sensing strategy was developed. Utilizing the corresponding aptamer of the antibiotics, strain displacement reactions were triggered, disrupting the ECL quenching system composed of perovskite and Ag nanoclusters (Ag NCs) on the electrode surface, generating a signal to achieve quantitative detection of several common tetracycline antibiotics. The perovskite quantum dot provided a strong and stable initial signal, while the efficient catalytic activity of the silver cluster enhanced the recognition sensitivity. Tetracycline, chlortetracycline, and oxytetracycline were used as examples to demonstrate the differentiation and quantitative detection through this method. In addition, the aptasensor exhibited analytical performance with the linear range (0.1-10 µM OTC) and good recovery rates of 94.7% to 101.6% in real samples. This approach has the potential to become a sensitive and practical approach for assessing antibiotic residues.
Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Compuestos de Calcio , Nanopartículas del Metal , Óxidos , Titanio , Tetraciclina , Técnicas Electroquímicas/métodos , Mediciones Luminiscentes/métodos , Antibacterianos , Tetraciclinas , Técnicas Biosensibles/métodos , Nanopartículas del Metal/química , Aptámeros de Nucleótidos/químicaRESUMEN
As a crucial indicator in food and water safety testing, the detection of Escherichia coli plays a significant role in maintaining environmental sanitation and promoting public health. Herein, based on the electrochemical activity characteristics of E. coli, we established an enhanced electrochemiluminescence aptasensor for E. coli analysis. This study presents a new method for accurate identification by utilizing a double aptamer recognition system. Specifically, a nano-cadmium sulfide (CdS) modified aptamer was used for primary labeling, while a second aptamer was immobilized on a graphene/chitosan composite electrode for re-capture. The use of two aptamers improves the accuracy of the identification process. Furthermore, the application of an electrode potential facilitates continuous electron transfer between the electrode and electrochemically active microorganisms, resulting in an enhanced electroluminescence signal in relation to the metabolic status. This strategy possesses better sensitivity, accuracy, and stability, demonstrating its potential for E. coli analysis.
Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Grafito , Escherichia coli/metabolismo , Técnicas Electroquímicas/métodos , Electrones , Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Grafito/químicaRESUMEN
Helicobacter pylori is closely linked to many gastric diseases such as gastric ulcers and duodenal ulcers. Therefore, biosensing H. pylori has attracted wide attention from both scientists and clinicians. Here, we proposed an electrochemiluminescence (ECL)-based platform that could sensitively detect H. pylori DNA. In this platform, a novel target-cycling synchronized rolling circle amplification was used for signal amplification. Silver nanoclusters (Ag NCs) were synthesized on the circle DNA products, embedding them with the ability to catalyze the electrochemical reduction of K2 S2 O8 , in turn resulting in rapid consumption of the ECL co-reactant near the working electrode, and leading to a decrease in the ECL emission intensity. In addition to its excellent stability and selectivity, the proposed strategy had a low detection limit of 10 pM, an indication that it can be beneficially applied to test biosamples. Furthermore, a biosensing chip was designed to improve the throughput and shed new light on large-scale clinical biosensing applications.
Asunto(s)
Técnicas Biosensibles , Helicobacter pylori , Nanopartículas del Metal , Helicobacter pylori/genética , Mediciones Luminiscentes/métodos , Técnicas Electroquímicas/métodos , Técnicas Biosensibles/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , ADN/genética , Límite de DetecciónRESUMEN
CRISPR/Cas9 has become a powerful tool for genomic manipulation, and the evaluation of the Cas9 activity is essential for the precise control of the CRISPR system. Herein, we develop a point-of-care platform for the rapid and visible determination of the Cas9 activity. A bipolar electrochemiluminescence (ECL) biosensor platform comprises an Au electric circuit fabricated on a glass substrate and the PDMS layer acts as a reservoir. DNA probes are then modified on electrodes and hybridization chain reaction (HCR) is applied to introduce ferrocene-labeled DNA. The CRISPR/Cas9 system can recognize the immobilized DNA probes and cleave them from the electrode surface as well as the labeled H1 and H2. Therefore, the cleavage activity is closely related to the labeled ferrocene. The modified electrode is then embodied in a closed bipolar system such that the oxidation reaction on sensing poles could be detected by the anodic ECL reactions on isolated luminescent poles due to charge balance, leading to the changes in ECL signals. This portable, integrated and convenient platform may become a meaningful tool for the discovery of a new CRISPR/Cas9 system or additional Cas9 inhibitors.
Asunto(s)
Técnicas Biosensibles/instrumentación , Proteína 9 Asociada a CRISPR/metabolismo , Electroquímica , Mediciones Luminiscentes , Secuencia de Bases , Sondas de ADN/genética , Sondas de ADN/metabolismo , ElectrodosRESUMEN
In this study, we developed a novel simple fluorescence resonance-energy transfer (FRET) system between two-color CdTe quantum dots (QDs) assisted by cetyltrimethylammonium bromide (CTAB). Mercaptopropionic (MPA)-capped CdTe QDs serving as both donors and acceptors were successfully synthesized by changing the refluxing time in aqueous solution. CTAB micelles formed in water and minimized the distance between the donors and acceptors significantly by electrostatic interactions, improving FRET efficiency. Several factors that affected the fluorescence spectra of the FRET system were investigated. The prepared FRET system was feasible as an effective fluorescent probe to detect Hg(II) in aqueous solution. At pH 7.0, a linear relationship between the quenched fluorescence intensity of orange-emitting acceptors (QDs(A) ) and Hg(II) concentration was acquired in the range 5-250 nmol/L with a detection limit of 1.95 nmol/L. The developed method showed excellent analytical performance for Hg(II) with high sensitivity and acceptable selectivity, reproducibility and stability. This finding indicated that the method has a promising potential application for environmental monitoring. This study demonstrated the great promise of QDs for expedient, low-cost and high-sensitivity detection of Hg(II).
Asunto(s)
Compuestos de Cadmio/química , Cetrimonio/química , Color , Transferencia Resonante de Energía de Fluorescencia , Mercurio/análisis , Puntos Cuánticos/química , Telurio/químicaRESUMEN
Though microfluidic paper analytical devices (µPADs) have attracted paramounting attentions in recent years as promising devices for low cost point-of-care tests, their real applications for blood analysis are still challenged by integrating sample preparation with different detection modes on a same µPAD. Herein, we developed a novel µPAD, which well coupled automatic serum extraction with reliable dual mode iron health tests: fluorescent analysis for Fe3+ and colorimetric ELISA for ferritin. All these functions are made available by in situ carbon dots (CDs) and AuNPs sequential patterning techniques. For CDs immobilization, hydrothermal reaction was taken on paper, to which a patterned through-hole polydimethylsiloxane (PDMS) mask was applied. None fluorescence CDs (nF-CDs) were generated on exposed regions, while the fluorescent CDs (F-CDs) were generated simultaneously on covered regions. Sensitive serum iron quantification was realized on the F-CDs modified regions, where Fe3+ ion can selectively quench the fluorescence of F-CDs. For AuNPs immobilization, electroless plating was taken on nF-CDs modified regions. The resulting AuNPs on nF-CDs layer on one hand triggered the coagulation of blood cells and thus led to the longest ever wicking distance for serum separation, on the other hand facilitated colorimetric enzyme linked immunosorbent assay (ELISA) for detection of serum ferritin. Combining the two readings, the µPAD can provide reliable measurement for serum iron and serum ferritin in whole blood. Furthermore, as CDs and AuNPs modified µPAD has the features of easy handling, low-cost, lightweight, and disposability, it is accounting for a promising prototype for whole blood point-of-care analysis.
Asunto(s)
Carbono/química , Compuestos Férricos/sangre , Ferritinas/sangre , Dispositivos Laboratorio en un Chip , Papel , Coagulación Sanguínea , Electrodos , Ensayo de Inmunoadsorción Enzimática , Fluorescencia , Oro/química , Humanos , Nanopartículas del Metal/química , Pruebas en el Punto de AtenciónRESUMEN
The detection of hydrogen peroxide (H2O2) represents an extensive requirement across various domains, including food, environmental, and medical fields. This study introduces a highly sensitive technique for the quantification of H2O2, integrating the electrochemiluminescence properties of perovskite with bio-catalyzed precipitation. A water-soluble perovskite-based electrochemiluminescence (ECL) biosensing interface was constructed, wherein H2O2 catalyzes a precipitation reaction that leads to the formation of an insoluble precipitate on the electrode surface. This occurrence effectively quenches the electrochemiluminescence signal of the perovskite, thus facilitating the quantitative detection of H2O2. The modified perovskite demonstrated excellent ECL performance, offering a stable signal source, while the bio-catalyzed precipitation reaction significantly amplified the quenching effect, thereby enhancing detection sensitivity. This strategy exhibits excellent stability and sensitivity, presenting a promising method for the detection of hydrogen peroxide, which holds great potential for applications in various fields.
RESUMEN
Helicobacter pylori infection has become a threat to the world populations. This leads to an urgent need of an efficient and convenient approach to accurately diagnose H. pylori infection. Saliva-based diagnoses are particularly welcomed for their efficiency and convenience. Aiming at saliva sample analysis, we proposed a CRISPR/Cas12a-assisted array, which had integrated H. pylori concentration detection and genotype screening functions. Single-nucleotide variations (SNVs) could be distinguished using the screening array with different probes, and an isothermal cycling strategy was combined with the trans-cleavage activity of Cas12a for signal amplification to improve accuracy of the diagnosis. As a demonstration, the SNV screening array was fabricated by utilizing the hybridization efficiency difference caused by mismatched bases. The array was able to successfully distinguish between ten H. pylori genotypes, and combined with the successful SDA biosensing, it had a LOD of as low as 60 fM. It was also able to diagnose H. pylori infection in saliva samples from infected patients. Together, the developed array has a potential in large-scale clinical screening and is a promising tool for the diagnosis and prevention of H. pylori infection-related diseases.
Asunto(s)
Técnicas Biosensibles , Infecciones por Helicobacter , Helicobacter pylori , Humanos , Saliva , Helicobacter pylori/genética , Infecciones por Helicobacter/diagnóstico , Infecciones por Helicobacter/genética , Sistemas CRISPR-Cas , ADNRESUMEN
Helicobacter pylori infections are threats to public health due to their high infection rate and drug resistance. Identification of single-nucleotide variants (SNVs) in H. pylori is crucial for both diagnosis and therapy. Yet the clinical testing of resistant H. pylori mutants is still facing some challenges, such as the selectivity is not good enough for SNVs in abundant wild-type DNA, the lack of clinical validation and the economical burden on patients. Herein, an X-shaped DNA probe with a toehold initiator was designed, which could specifically hybridize with certain genotype DNA due to the thermodynamically driven reaction. A competitive reaction was developed to amplify the thermodynamic difference between wild-type DNA and SNVs, diminishing the interference of wild-type DNA. By this means, multiple SNVs in H. pylori were successfully identified and two SNVs related to clarithromycin resistance are chosen as model targets. A paper strip was fabricated for visual, fast screening of SNVs. Furthermore, the approach was validated using clinical samples, and a point-of-care (POCT) testing diagnosis was executed on saliva samples, demonstrating its potential for the prevention and cure of H. pylori infections.
Asunto(s)
Infecciones por Helicobacter , Helicobacter pylori , Antibacterianos/farmacología , Claritromicina/farmacología , Claritromicina/uso terapéutico , Farmacorresistencia Bacteriana/genética , Infecciones por Helicobacter/diagnóstico , Infecciones por Helicobacter/tratamiento farmacológico , Helicobacter pylori/genética , Humanos , Pruebas de Sensibilidad Microbiana , NucleótidosRESUMEN
Metal ion synergistically enhanced chemotherapy is a promising strategy for cancer treatment. However, targeting delivery of ions toward cancer cells remains challenging for decades. Herein, we developed a novel folic acid-nanogel (termed as FA-nanogel) with alkane chains as diffusion barriers for targeted transmembrane delivery of calcium ions into cancer cells. With the aid of hydrophobic diffusion barriers, the FA-nanogel showed a reduced and sustained speed for release of calcium ions, significantly prolonging the ion effect. Moreover, a pH-sensitive injectable hydrogel-loaded FA-nanogel and chemotherapeutic drug 5-fluorouracil (5-Fu) was synthesized for investigating the synergistic effect of nanogel on chemotherapy. Both in vitro and in vivo experiments confirmed that the intracellular calcium ions were continuously increased because of the targeted delivery ability and ion sustained release ability of the smart FA-nanogel, and the tumor growth was effectively inhibited by the ion synergistic chemotherapy. This study not only provides a powerful nanoplatform for sustained transmembrane delivery of ions into malignant cells but also creates better conditions for improving the therapeutic efficacy of chemotherapy.
Asunto(s)
Calcio/administración & dosificación , Hidrogeles/administración & dosificación , Nanopartículas del Metal/administración & dosificación , Neoplasias/tratamiento farmacológico , Animales , Calcio/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Doxorrubicina/administración & dosificación , Doxorrubicina/química , Portadores de Fármacos/administración & dosificación , Portadores de Fármacos/química , Fluorouracilo/administración & dosificación , Fluorouracilo/química , Ácido Fólico/administración & dosificación , Ácido Fólico/química , Células HeLa , Humanos , Hidrogeles/síntesis química , Hidrogeles/química , Concentración de Iones de Hidrógeno , Iones/administración & dosificación , Iones/química , Nanopartículas del Metal/química , Ratones , Neoplasias/patología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The development of non-invasive techniques for the diagnosis of cancer, characterization of mutation and monitoring treatment response could greatly reduce the morbidity and mortality caused by cancer. Nevertheless, the extremely low amount of cell free nucleic acids makes liquid biopsy a very challenging task. Herein, taking advantage of the pocket size, reliable quantitative results and simple operation of the pocket-sized personal glucose meter (PGM), we report an approach of circulating microRNA-21 (miR-21) detection with high precision and low cost. Via target-induced release of invertase from the DNA-invertase conjugate, which could convert sucrose into glucose, the detection of miR-21 in serum was linked to PGM readings. Combining the DNAzyme feedback amplification (DFA) program and highly efficient enzymatic turnover, an ultralow detection limit of 7 × 10-16 M for miR-21 was achieved using a PGM as the reporter. The high sensitivity and selectivity of the proposed method meets the requirement of quantifying cell free nucleic acids in serum. In addition, this approach fills the shortage of quantitative RT-PCR and next-generation sequencing in quantifying miRNAs with a short length and greatly reduces the cost of detection. We believe that widely used personal diagnosis devices could hold an important place in the booming area of liquid biopsy.
RESUMEN
A paper-based SERS test strip combining strengths of paper chip and Raman active substrate was demonstrated to overcome challenges in spectroscopic sensing of complicated samples and realize quantitative detection of disease markers in whole blood. The precisely controlled Au NPs were not only capable of generating condensed hot spots on the fibers, but also enhanced the size exclusion effect of paper, resulting in the novel performance on both SERS detection and sample pretreatment. A biosensor for Mucin-1 is developed by equipping the Au NPs with aptamer. Combining all these merits, this small, cheap and portable test strip might find wide application in clinical diagnosis and health evaluation.
Asunto(s)
Técnicas Biosensibles , Oro/química , Nanopartículas del Metal/química , Mucina-1/sangre , Espectrometría Raman/métodos , Aptámeros de Nucleótidos/síntesis química , Aptámeros de Nucleótidos/química , Carbocianinas , Colorantes Fluorescentes , Humanos , Tiras Reactivas/químicaRESUMEN
In this paper, a novel prototype liquid-air dual gradient chip is introduced, which has paved the way for effective synergic effect bio-evaluation of air pollutant. The chip is composed of an array of the agarose liquid-air interfaces, top air gradient layer and bottom liquid gradient layer. The novel agarose liquid-air interface allows for non-biased exposure of cells to all the substances in the air and diffusive interactions with the liquid phase; while the dual liquid-air gradient provides powerful screening abilities, which well reduced errors, saved time and cost from repeated experiment. Coupling the two functions, the chip subsequently facilitates synergic effect evaluation of both liquid and air factors on cells. Here cigarette smoke was taken as the model air pollutant, and its strong synergic effects with inflammatory level of A549 lung cancer cells on their fate were successfully quantified for the first time. These results well testified that the proposed dual-gradient chip is powerful and indispensable for bio-evaluation of air pollutant.
Asunto(s)
Contaminantes Atmosféricos/toxicidad , Dispositivos Laboratorio en un Chip , Humo/análisis , Contaminación por Humo de Tabaco/análisis , Células A549 , Aire/análisis , Apoptosis/efectos de los fármacos , Diseño de Equipo , Humanos , Necrosis/inducido químicamente , Perfusión , Sefarosa/química , Agua/químicaRESUMEN
In this work, we report a novel microfluidic gas collecting platform aiming at simultaneous sample extraction and multiplex mass spectrometry (MS) analysis. An alveolar-mimicking elastic polydimethylsiloxane (PDMS) structures was designed to move dynamically driven by external pressure. The movement was well tuned both by its amplitude and rhythm following the natural process of human respiration. By integrating the alveolar units into arrays and assembling them to gas channels, a cyclic contraction/expansion system for gas inhale and exhale was successfully constructed. Upon equipping this system with a droplet array on the alveolar array surface, we were able to get information of inhaled smoke in a new strategy. Here, with cigarette smoke as an example, analysis of accumulation for target molecules during passive smoking is taken. Relationships between the breathing times, distances away from smokers and inhaled content of nicotine are clarified. Further, by applying different types of extraction solvent droplets on different locations of the droplet array, simultaneous extraction of nicotine, formaldehyde and caproic acid in sidestream smoke (SS) are realized. Since the extract droplets are spatially separated, they can be directly analyzed by MS which is fast and can rid us of all complex sample separation and purification steps. Combining all these merits, this small, cheap and portable platform might find wide application in inhaled air pollutant analysis both in and outdoors.
Asunto(s)
Contaminantes Atmosféricos/análisis , Contaminantes Atmosféricos/aislamiento & purificación , Fraccionamiento Químico/instrumentación , Dispositivos Laboratorio en un Chip , Humo/análisis , Productos de Tabaco/análisis , Biomimética , Pruebas Respiratorias , Monóxido de Carbono/análisis , Monóxido de Carbono/aislamiento & purificación , Dimetilpolisiloxanos/química , Humanos , Nicotina/análisis , Nicotina/aislamiento & purificación , Contaminación por Humo de Tabaco/análisisRESUMEN
We propose a novel method to obtain versatile droplets arrays on a regional hydrophilic chip that is fabricated by PDMS soft lithography and regional plasma treatment. It enables rapid liquid dispensation and droplets array formation just making the chip surface in contact with solution. By combining this chip with a special Christmas Tree structure, the droplets array with concentrations in gradient is generated. It possesses the greatly improved performance of convenience and versatility in bioscreening and biosensing. For example, high throughput condition screening of toxic tests of CdSe quantum dots on HL-60 cells are conducted and cell death rates are successfully counted quickly and efficiently. Furthermore, a rapid biosensing approach for cancer biomarkers carcinoma embryonic antigen (CEA) is developed via magnetic beads (MBs)-based sandwich immunoassay methods.
Asunto(s)
Técnicas Analíticas Microfluídicas , Microfluídica/métodos , Técnicas Biosensibles , Compuestos de Cadmio/química , Antígeno Carcinoembrionario/química , Dimetilpolisiloxanos/química , Elastómeros/química , Células HL-60 , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Inmunoensayo , Magnetismo , Ensayo de Materiales , Nanopartículas/química , Nanotecnología/métodos , Puntos Cuánticos , Compuestos de Selenio/química , Propiedades de Superficie , HumectabilidadRESUMEN
Based on the ingenious combination of two different gradient generation mechanisms, this work reports a novel approach for a high throughput linear liquid gradient in a two-dimensional (2D) matrix. Specifically, a typical Christmas Tree structure with two inlets was designed as the first mixture gradient generator, upon which the second diffusion gradient generator was coupled to produce the desired concentration series on the basis of the distance difference. Rather than a simple 1D line, the integration of the two generators would result in an innovative 2D matrix of reservoirs, which was then characterized both theoretically and experimentally. Theoretically, calculation of fluid field demonstrated the formation of a concentration gradient, which was then confirmed by the dye solution visualization analysis. For high throughput screening application, doxorubicin (Dox) was then selected as model medicine to treat the acute myeloblastic leukemia (HL-60) cells. Cell viability displayed that cell death rate enhanced with the increase of drug concentration, and this result was higher than that on a 96-well plate, and the corresponding mechanism was properly discussed. Subsequently, Dox and quercetin were employed simultaneously to generate an overlapping gradient and its effect on HL-60 cells was investigated. Due to the automatic formation of concentration gradient that could improve the work efficiency, this work provides a promising tool for future high throughput drug screening.
RESUMEN
In this work a novel microfluidic platform for cell culture and assay is developed. On the chip a static cell culture region is coupled with dynamic fluidic nutrition supply structures. The cell culture unit has a sandwich structure with liquid channels on the top, the cell culture reservoir in the middle and gas channels on the bottom. Samples can be easily loaded into the reservoir and exchange constantly with the external liquid environment by diffusion. Since the flow direction is perpendicular to the liquid channel on the top of the reservoir, the cells in the reservoir are shielded from shear-force. By assembling the basic units into an array, a steady concentration gradient can be generated. Cell culture models both for continuous perfusion and one-off perfusion were established on the chip. Both adherent and suspended cells were successfully cultured on the chip in 2D and 3D culture modes. After culturing, the trapped cells were recovered for use in a later assay. As a competitive candidate for a standard cell culture and assay platform, this chip is also adaptable for cytotoxicity and cell growth assays.
Asunto(s)
Técnicas de Cultivo de Célula/instrumentación , Técnicas Analíticas Microfluídicas/instrumentación , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Dimetilpolisiloxanos/química , Vidrio/química , Células HL-60 , Humanos , Nanopartículas del Metal/química , Nanopartículas del Metal/toxicidad , Nanotubos de Carbono/química , Nanotubos de Carbono/toxicidad , Compuestos de Estaño/químicaRESUMEN
In this study, steady liquid-gas phase separation is realized by applying a hydrophobic small microchannel array (SMA) to bridge two large microchannels, one for liquid phase and one for gas phase. In this structure, a capillary pressure difference between that in the SMA and the larger channel results in a steady liquid-gas interface. The generated liquid-gas interface allows for fast gas dissolving speed. By coupling the liquid-gas interface with a one directional fluidic field, a steady dissolved gas concentration gradient (DgCG) is generated. The DgCG distribution is easily designable for linear or exponential modes, providing improved flexibility for gas participated processes on chip. To demonstrate its applicability, a CO(2) DgCG chip is fabricated and applied for screening CaCO(3) crystal growth conditions in the DgCG chip. Crystals with transitional structures are successfully fabricated, which is consistent with the CO(2) DgCG distribution.