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1.
Anal Chem ; 96(21): 8641-8647, 2024 05 28.
Artículo en Inglés | MEDLINE | ID: mdl-38716697

RESUMEN

Pathogenic bacterial infections, even at extremely low concentrations, pose significant threats to human health. However, the challenge persists in achieving high-sensitivity bacterial detection, particularly in complex samples. Herein, we present a novel sandwich-type electrochemical sensor utilizing bacteria-imprinted polymer (BIP) coupled with vancomycin-conjugated MnO2 nanozyme (Van@BSA-MnO2) for the ultrasensitive detection of pathogenic bacteria, exemplified by Staphylococcus aureus (S. aureus). The BIP, in situ prepared on the electrode surface, acts as a highly specific capture probe by replicating the surface features of S. aureus. Vancomycin (Van), known for its affinity to bacterial cell walls, is conjugated with a Bovine serum albumin (BSA)-templated MnO2 nanozyme through EDC/NHS chemistry. The resulting Van@BSA-MnO2 complex, serving as a detection probe, provides an efficient catalytic platform for signal amplification. Upon binding with the captured S. aureus, the Van@BSA-MnO2 complex catalyzes a substrate reaction, generating a current signal proportional to the target bacterial concentration. The sensor displays remarkable sensitivity, capable of detecting a single bacterial cell in a phosphate buffer solution. Even in complex milk matrices, it maintains outstanding performance, identifying S. aureus at concentrations as low as 10 CFU mL-1 without requiring intricate sample pretreatment. Moreover, the sensor demonstrates excellent selectivity, particularly in distinguishing target S. aureus from interfering bacteria of the same genus at concentrations 100-fold higher. This innovative method, employing entirely synthetic materials, provides a versatile and low-cost detection platform for Gram-positive bacteria. In comparison to existing nanozyme-based bacterial sensors with biological recognition materials, our assay offers distinct advantages, including enhanced sensitivity, ease of preparation, and cost-effectiveness, thereby holding significant promise for applications in food safety and environmental monitoring.


Asunto(s)
Compuestos de Manganeso , Óxidos , Polímeros , Staphylococcus aureus , Vancomicina , Staphylococcus aureus/aislamiento & purificación , Compuestos de Manganeso/química , Óxidos/química , Vancomicina/química , Polímeros/química , Albúmina Sérica Bovina/química , Técnicas Electroquímicas/métodos , Análisis de la Célula Individual , Antibacterianos/química , Antibacterianos/farmacología , Animales , Límite de Detección , Polímeros Impresos Molecularmente/química , Humanos
2.
Analyst ; 148(14): 3239-3246, 2023 Jul 10.
Artículo en Inglés | MEDLINE | ID: mdl-37341575

RESUMEN

Microfluidic impedance cytometry is emerging as a label-free, low-cost and portable solution for cell analysis. Impedance-based cell or particle characterization is provided by microfluidic and electronic devices. We report the design and characterization of a miniaturized flow cytometer based on a 3-dimensional (3D) hydrodynamic focusing mechanism. A sheath adaptively concentrated the sample laterally and vertically at the bottom of the microchannel, reducing the variance of particle translocation height and increasing the signal-to-noise ratio of the particle impedance pulse. Through simulation and confocal microscopy experiments, it has been verified that an increase in the ratio of sheath to sample decreased the cross-sectional area of the concentrated stream, which can be reduced to 26.50% of the pre-focusing value. The appropriate sheath flow settings increased the impedance pulse amplitude for different particles, and the coefficient of variation reduced by at least 35.85%, contributing to a more accurate representation of the particle impedance characteristic distribution. The system displayed the difference of HepG2 cell impedance before and after drug treatment, which is consistent with the results of flow cytometry, providing a convenient and inexpensive solution for monitoring cell status.


Asunto(s)
Técnicas Analíticas Microfluídicas , Microfluídica , Técnicas Analíticas Microfluídicas/métodos , Citometría de Flujo/métodos , Impedancia Eléctrica , Hidrodinámica
3.
Analyst ; 148(12): 2758-2766, 2023 Jun 12.
Artículo en Inglés | MEDLINE | ID: mdl-37194305

RESUMEN

This paper introduces an enclosed microfluidic chip that integrates sample preparation and the chamber-based digital polymerase chain reaction (cdPCR). The sample preparation of the chip includes nucleic acid extraction and purification based on magnetic beads, which adsorb nucleic acids by moving around the reaction chambers to complete the reactions including lysis, washing, and elution. The cdPCR area of the chip consists of tens of thousands of regularly arranged microchambers. After the sample preparation processes are completed, the purified nucleic acid can be directly introduced into the microchambers for amplification and detection on the chip. The nucleic acid extraction performance and digital quantification performance of the system were examined using synthetic SARS-CoV-2 plasmid templates at concentrations ranging from 101-105 copies per µL. Further on, a simulated clinical sample was used to test the system, and the integrated chip was able to accurately detect SARS-CoV-2 virus particle samples doped with interference (saliva) with a detection limit of 10 copies per µL. This integrated system could provide a promising tool for point-of-care testing of pathogenic infections.


Asunto(s)
Microfluídica , Microfluídica/métodos , Reacción en Cadena de la Polimerasa , Ácidos Nucleicos/análisis , SARS-CoV-2/genética , SARS-CoV-2/aislamiento & purificación
4.
Analyst ; 148(9): 1939-1947, 2023 May 02.
Artículo en Inglés | MEDLINE | ID: mdl-36916483

RESUMEN

Diagnosis of cancer by biomarkers plays an important role in human health and life. However, current laboratory techniques for detecting cancer biomarkers still require laborious and time-consuming operation by skilled operators and associated laboratory instruments. This work presents a colorimetric biosensor for the rapid and sensitive detection of carcinoembryonic antigen (CEA) based on an automated immunomagnetic separation platform and a droplet array microfluidic chip with the aid of an image analysis system. Immunomagnetic nanoparticles (MNPs) were used to capture CEA in the samples. CEA-detecting antibodies and horseradish peroxidase (HRP) were modified on polystyrene microspheres (PS), catalysing hydrogen peroxide and 3,3',5,5'-tetramethylbenzidine (TMB) as signal outputs. Color reaction data were analyzed to establish a CEA concentration standard curve. The movement of MNPs between droplets in the microfluidic chip is achieved using an automatically programmable magnetic control system. This colorimetric biosensor has been used for the simultaneous detection of six CEA samples ranging from 100 pg mL-1 to 100 ng mL-1 with a detection limit of 14.347 pg mL-1 in 10 min, following the linear equation: y = -4.773 ln(x) + 156.26 with a correlation of R2 = 0.9924, and the entire workflow can be completed within 80 minutes. The microfluidic immunosensor designed in this paper has the advantages of low cost, automation, low sample consumption, high throughput, and promising applications in biochemistry.


Asunto(s)
Técnicas Biosensibles , Nanopartículas del Metal , Humanos , Antígeno Carcinoembrionario/análisis , Separación Inmunomagnética/métodos , Microfluídica , Inmunoensayo/métodos , Técnicas Biosensibles/métodos , Anticuerpos Monoclonales , Límite de Detección , Oro
5.
BMC Plant Biol ; 22(1): 596, 2022 Dec 19.
Artículo en Inglés | MEDLINE | ID: mdl-36536303

RESUMEN

BACKGROUND: Late embryogenesis abundant (LEA) proteins are widely distributed in higher plants and play crucial roles in regulating plant growth and development processes and resisting abiotic stress. Cultivated tomato (Solanum lycopersicum) is an important vegetable crop worldwide; however, its growth, development, yield, and quality are currently severely constrained by abiotic stressors. In contrast, wild tomato species are more tolerant to abiotic stress and can grow normally in extreme environments. The main objective of this study was to identify, characterize, and perform gene expression analysis of LEA protein families from cultivated and wild tomato species to mine candidate genes and determine their potential role in abiotic stress tolerance in tomatoes. RESULTS: Total 60, 69, 65, and 60 LEA genes were identified in S. lycopersicum, Solanum pimpinellifolium, Solanum pennellii, and Solanum lycopersicoides, respectively. Characterization results showed that these genes could be divided into eight clusters, with the LEA_2 cluster having the most members. Most LEA genes had few introns and were non-randomly distributed on chromosomes; the promoter regions contained numerous cis-acting regulatory elements related to abiotic stress tolerance and phytohormone responses. Evolutionary analysis showed that LEA genes were highly conserved and that the segmental duplication event played an important role in evolution of the LEA gene family. Transcription and expression pattern analyses revealed different regulatory patterns of LEA genes between cultivated and wild tomato species under normal conditions. Certain S. lycopersicum LEA (SlLEA) genes showed similar expression patterns and played specific roles under different abiotic stress and phytohormone treatments. Gene ontology and protein interaction analyses showed that most LEA genes acted in response to abiotic stimuli and water deficit. Five SlLEA proteins were found to interact with 11 S. lycopersicum WRKY proteins involved in development or resistance to stress. Virus-induced gene silencing of SlLEA6 affected the antioxidant and reactive oxygen species defense systems, increased the degree of cellular damage, and reduced drought resistance in S. lycopersicum. CONCLUSION: These findings provide comprehensive information on LEA proteins in cultivated and wild tomato species and their possible functions under different abiotic and phytohormone stresses. The study systematically broadens our current understanding of LEA proteins and candidate genes and provides a theoretical basis for future functional studies aimed at improving stress resistance in tomato.


Asunto(s)
Solanum lycopersicum , Solanum , Reguladores del Crecimiento de las Plantas , Sequías , Proteínas de Plantas/genética , Perfilación de la Expresión Génica , Solanum/genética , Estrés Fisiológico/genética , Regulación de la Expresión Génica de las Plantas , Filogenia
6.
Anal Biochem ; 656: 114877, 2022 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-36055398

RESUMEN

The lack of reliable and practical method for detecting rare hot mutation of epidermal growth factor receptor (EGFR) in circulating tumor DNA (ctDNA) for lung cancer has remained a challenge for general clinical application due to excess wild type DNA in clinical samples. In this study, we developed a droplet digital PCR (ddPCR) platform, integrating a PDMS chip and double-layer glass reservoir. The duplex T-junction droplet generators in PDMS chip can produce about one million uniform droplets of 4.187 pL within ∼10 min, which were then stored in the glass reservoir. The double-layer glass reservoir can protect droplets from evaporation and breaking, solving the problem of instability during thermal-cycling. The quantitative capabilities of the ddPCR chip were evaluated by testing EGFR exon gene 21, with a good linear correlation in the wide range of 101 to 106 copies/µL (R2 = 0.9998). We then demonstrated that the proposed ddPCR device can recognize rare EGFR L858R mutation under a background of 106 copies/µL wild-type DNA at a sensitivity of 0.0001%. Finally, we demonstrated this ddPCR platform could identify low amount of EGFR L858R mutation in ctDNA and CTCs of patients with lung cancer.


Asunto(s)
ADN Tumoral Circulante , Neoplasias Pulmonares , ADN Tumoral Circulante/genética , Receptores ErbB/genética , Genes erbB-1 , Humanos , Neoplasias Pulmonares/diagnóstico , Mutación , Reacción en Cadena de la Polimerasa/métodos
7.
Biomed Microdevices ; 22(1): 18, 2020 02 15.
Artículo en Inglés | MEDLINE | ID: mdl-32076843

RESUMEN

Centrifugal droplet-based microfluidic devices have been applied to biomedical analysis and diagnostics recently. However, in centrifugal droplet-based microfluidic devices, droplets are tightly packed (i.e., the oil film between neighbouring droplets is thin). Therefore, droplet coalescence usually occurs especially during thermal incubation process. To preserve individual droplets in the devices, we report a new design for monodisperse droplet generation and storage that exploits a centrifugal configuration for droplet emulsification and oil-storage structures (OSSs) for regulation of the thickness of oil film between neighbouring droplets. The centrifugal emulsifier was well designed to ensure uniform droplet generation. Meanwhile, the OSSs could store oil during centrifugal emulsification while release oil before thermal incubation, which "loosen" tightly packed droplets to prevent droplets from coalescing. In this paper, the working process of OSS was analysed, and its shape and size were optimized. Then, the optimized OSSs were integrated into a centrifugal emulsifier for droplet digital loop mediated isothermal amplification (ddLAMP) by which detection of JAK2 V617F mutation within myeloproliferative neoplasms with a dynamic range of 101 to 104 copies per µL was achieved. We anticipate that the simplicity and robustness of our system make it attractive as an inexpensive and easy-to-operate device for DNA amplification, particularly applicable in point-of-care settings.


Asunto(s)
Dispositivos Laboratorio en un Chip , Técnicas Analíticas Microfluídicas , Técnicas de Diagnóstico Molecular , Técnicas de Amplificación de Ácido Nucleico , Aceites/química , Sustitución de Aminoácidos , Centrifugación , Emulsiones , Neoplasias Hematológicas/genética , Humanos , Janus Quinasa 2/genética , Mutación Missense , Trastornos Mieloproliferativos/genética , Proteínas de Neoplasias/genética
9.
Langmuir ; 32(48): 12623-12631, 2016 12 06.
Artículo en Inglés | MEDLINE | ID: mdl-27934532

RESUMEN

Two-dimensional graphene devices are widely used for biomolecule detection. Nevertheless, the surface modification of graphene is critical to achieve the high sensitivity and specificity required for biological detection. Herein, native bovine serum albumin (BSA) in inorganic solution is denatured on the graphene surface by heating, leading to the formation of nanoscale BSA protein films adsorbed on the graphene substrate via π-stacking interactions. This technique yields a controllable, scalable, uniform, and high-coverage method for graphene biosensors. Further, the application of such nanoscale heat-denatured BSA films on graphene as a universal graphene biosensor platform is explored. The thickness of heat-denatured BSA films increased with heating time and BSA concentration but decreased with solvent concentration as confirmed by atomic force microscopy. The noncovalent interaction between denatured BSA films and graphene was investigated by Raman spectroscopy. BSA can act as a p-type and n-type dopant by modulating pH-dependent net charges on the layered BSA-graphene surface, as assessed by current-voltage measurements. Chemical groups of denatured BSA films, including amino and carboxyl groups, were verified by X-ray photoelectron microscopy, attenuated total reflectance-Fourier transform infrared spectra, and fluorescent labeling. The tailoring of the BSA-graphene surfaces through chemical modification, controlled thickness, and doping type via noncovalent interactions provides a controllable, multifunctional biosensor platform for molecular diagnosis without the possibility of nonspecific adsorption on graphene.


Asunto(s)
Grafito/química , Albúmina Sérica Bovina/química , Adsorción , Animales , Técnicas Biosensibles , Bovinos , Colorantes Fluorescentes/química , Calor , Concentración de Iones de Hidrógeno , Microscopía de Fuerza Atómica , Nanoestructuras , Conformación Proteica , Desnaturalización Proteica , Solventes , Espectroscopía Infrarroja por Transformada de Fourier , Espectrometría Raman , Propiedades de Superficie
10.
J Clin Lab Anal ; 28(2): 104-9, 2014 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-24395581

RESUMEN

BACKGROUND: Urine protein test has been widely used in clinics, but to determine the type of proteinuria is usually difficult due to technical limitations. METHODS: In the current study, a rapid and simple method to separate and determine urine proteins by a microchip electrophoresis (ME) system has been developed in which only 4 min are required. RESULTS: Optimal separation conditions have been established by using 15 s injection time at 500 and 1,500 V separation voltage in 75 mmol/l borate buffer containing 0.8 mmol/l calcium lactate and 1% ϕ ethylamine (pH 10.55). Relative standard deviation (RSD) of migration time with purified human albumin and human transferring was 2.68% and 2.24%, and RSD of the peak area was 5.85% and 4.96%, respectively. The linear detection range was 1.0-15.0 g/l for purified human albumin and 1.0-10.0 g/l for human transferrin, with the same detection limit (S/N = 3) of 0.4 g/l. Finally, comparing to conventional agarose gel electrophoresis, the same results were obtained by using ME by testing clinical samples including 60 selective proteinuria, 105 nonselective proteinuria, and 6 overflow proteinuria. CONCLUSION: This newly established ME could have broad applications to determine the type of proteinuria in clinics.


Asunto(s)
Electroforesis por Microchip/métodos , Proteínas/aislamiento & purificación , Proteinuria/diagnóstico , Orina/química , Tampones (Química) , Compuestos de Calcio/química , Electricidad , Etilaminas/química , Humanos , Lactatos/química , Límite de Detección , Análisis de Regresión , Reproducibilidad de los Resultados , Soluciones , Factores de Tiempo
11.
Future Oncol ; 9(10): 1489-500, 2013 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-24106900

RESUMEN

AIM: The objective of this study was to investigate the clinical significance of circulating tumor cells (CTCs) on the evaluation and prediction of treatment responses in rectal cancer patients compared with serum carcinoembryonic antigen (CEA). MATERIALS & METHODS: Both CTCs and CEA levels of 103 rectal cancer patients (66 with stage II-III and 37 with recurrence or metastasis) were analyzed before and after chemoradiotherapy. CTCs were detected using EpCAM magnetic bead-based enrichment combined with cytometric identification. RESULTS: CTCs were detected in all patients while no tumor cells were found in healthy controls. CTC levels in metastatic patients were significantly higher than those with recurrence or stage II-III rectal cancer. There is a close relationship between CTC levels and treatment outcomes but serum CEA did not have any correlation. CONCLUSION: CTCs are promising markers for the evaluation and prediction of treatment responses in rectal cancer patients, superior to the conventional tumor marker CEA.


Asunto(s)
Antígeno Carcinoembrionario/sangre , Células Neoplásicas Circulantes/patología , Neoplasias del Recto/sangre , Neoplasias del Recto/patología , Adulto , Anciano , Línea Celular Tumoral , Progresión de la Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Metástasis de la Neoplasia , Estadificación de Neoplasias , Neoplasias del Recto/diagnóstico , Neoplasias del Recto/terapia , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Resultado del Tratamiento
12.
Talanta ; 265: 124776, 2023 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-37348357

RESUMEN

The isolation of single cell or droplet is first and crucial step to single-cell analysis, which is important for cancer research and diagnostic methods. This review provides an overview of technologies that are currently used or in development to realize the isolation. Microfluidic based manipulation is an emerging technology with the distinct advantages of miniaturization and low cost. Therefore, recent developments in microfluidic isolated methods have attracted extensive attention. We introduced herein five strategies based on microfluid: trap, microfluidic discrete manipulation, bioprinter, capillary and inertial force. For every technology, their basic principles and features were discussed firstly. Then some modified approaches and applications were listed as the extension. Finally, we compared the advantages and drawbacks of these methods, and analyzed the trend of the manipulation based on microfluidics.


Asunto(s)
Técnicas Analíticas Microfluídicas , Microfluídica , Microfluídica/métodos , Miniaturización , Análisis de la Célula Individual
13.
Biosensors (Basel) ; 13(5)2023 May 05.
Artículo en Inglés | MEDLINE | ID: mdl-37232879

RESUMEN

We developed a microfluidic chip integrated with nucleic acid purification and droplet-based digital polymerase chain reaction (ddPCR) modules to realize a 'sample-in, result-out' infectious virus diagnosis. The whole process involved pulling magnetic beads through drops in an oil-enclosed environment. The purified nucleic acids were dispensed into microdroplets by a concentric-ring, oil-water-mixing, flow-focusing droplets generator driven under negative pressure conditions. Microdroplets were generated with good uniformity (CV = 5.8%), adjustable diameters (50-200 µm), and controllable flow rates (0-0.3 µL/s). Further verification was provided by quantitative detection of plasmids. We observed a linear correlation of R2 = 0.9998 in the concentration range from 10 to 105 copies/µL. Finally, this chip was applied to quantify the nucleic acid concentrations of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The measured nucleic acid recovery rate of 75 ± 8.8% and detection limit of 10 copies/µL proved its on-chip purification and accurate detection abilities. This chip can potentially be a valuable tool in point-of-care testing.


Asunto(s)
COVID-19 , Ácidos Nucleicos , Humanos , SARS-CoV-2 , COVID-19/diagnóstico , Reacción en Cadena de la Polimerasa , Ácidos Nucleicos/análisis , Análisis de Secuencia por Matrices de Oligonucleótidos
14.
Anal Chim Acta ; 1271: 341469, 2023 Aug 29.
Artículo en Inglés | MEDLINE | ID: mdl-37328249

RESUMEN

Traditional nucleic acid extraction and detection is based on open operation, which may cause cross-contamination and aerosol formation. This study developed a droplet magnetic-controlled microfluidic chip integrated nucleic acid extraction, purification and amplification. The reagent is sealed in oil to form a droplet, and the nucleic acid is extracted and purified by controlling the movement of the magnetic beads (MBs) through a permanent magnet, ensuring a closed environment. This chip can automatically extract nucleic acid from multiple samples within 20 min, and can be directly placed in the in situ amplification instrument for amplification without further transfer of nucleic acid, characterized by simple, fast, time-saving and labor-saving. The results showed that the chip was able to detect <10 copies/test SARS-CoV-2 RNA, and EGFR exon 21 L858R mutations were detected in H1975 cells as low as 4 cells. In addition, on the basis of the droplet magnetic-controlled microfluidic chip, we further developed a multi-target detection chip, which used MBs to divide the nucleic acid of the sample into three parts. And the macrolides resistance mutations A2063G and A2064G, and the P1 gene of mycoplasma pneumoniae (MP) were successfully detected in clinical samples by the multi-target detection chip, providing the possibility for future application in the detection of multiple pathogens.


Asunto(s)
COVID-19 , Neoplasias , Ácidos Nucleicos , Humanos , Ácidos Nucleicos/genética , Microfluídica , ARN Viral , Técnicas de Amplificación de Ácido Nucleico/métodos , COVID-19/diagnóstico , SARS-CoV-2 , Fenómenos Magnéticos
15.
Nat Genet ; 55(5): 852-860, 2023 05.
Artículo en Inglés | MEDLINE | ID: mdl-37024581

RESUMEN

Effective utilization of wild relatives is key to overcoming challenges in genetic improvement of cultivated tomato, which has a narrow genetic basis; however, current efforts to decipher high-quality genomes for tomato wild species are insufficient. Here, we report chromosome-scale tomato genomes from nine wild species and two cultivated accessions, representative of Solanum section Lycopersicon, the tomato clade. Together with two previously released genomes, we elucidate the phylogeny of Lycopersicon and construct a section-wide gene repertoire. We reveal the landscape of structural variants and provide entry to the genomic diversity among tomato wild relatives, enabling the discovery of a wild tomato gene with the potential to increase yields of modern cultivated tomatoes. Construction of a graph-based genome enables structural-variant-based genome-wide association studies, identifying numerous signals associated with tomato flavor-related traits and fruit metabolites. The tomato super-pangenome resources will expedite biological studies and breeding of this globally important crop.


Asunto(s)
Solanum lycopersicum , Solanum , Solanum lycopersicum/genética , Estudio de Asociación del Genoma Completo , Genoma de Planta/genética , Fitomejoramiento , Solanum/genética , Genómica
16.
J Endourol ; 36(1): 143-150, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34098728

RESUMEN

Introduction: To evaluate the efficacy and safety of performing extracorporeal physical vibrational lithecbole (EPVL) through greater sciatic foramen (GSF) for distal ureteral calculi (DUC) treatment. Materials and Methods: All patients with a diagnosis of DUC (6-10 mm in diameter) were enrolled in this study from October 2018 to May 2020. Patients were randomly divided into three groups receiving EPVL through GSF (Group A, n = 58), or abdominal (Group B, n = 60), or combined with oral use of tamsulosin at 0.4 mg daily (Group C, n = 63). Results: There was no significant difference observed in terms of demographic characteristics or size of stones among the three groups (p > 0.05). Compared with the Groups B and C, patients of the Group A displayed a significantly higher score of comfort, with a significantly decreased number of renal colic attacks or analgesics required (p < 0.01). The stone-free rate also significantly increased after 1 and 2 weeks of treatment (p < 0.01), despite such a significant difference among these groups vanishing after 4 weeks of treatment. Conclusion: EPVL in the prone position uses the GSF as the path and is a safe and effective approach to treat the distal ureteral calculi.


Asunto(s)
Litotricia , Cálculos Ureterales , Humanos , Tamsulosina/uso terapéutico , Resultado del Tratamiento , Cálculos Ureterales/cirugía , Vibración
17.
Talanta ; 237: 122930, 2022 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-34736667

RESUMEN

Given increasing concern regarding antibiotic environmental contamination, there is immediate need to monitor antibiotic levels to effectively control pollution. In this study, we used a photoelectrochemical aptasensor based on TiO2@MoS2 spiral nanoarrays to detect chloramphenicol (CAP) in antibiotics. Nanoarrays were directly grown on fluorine-doped tin oxide (FTO) conductive glass with excellent biochemical stability, while aptamer-SH were immobilized by chemical binding on a synthetic TiO2@MoS2 nanoarray. Results show that the photocurrents were reduced in the presence of photoelectrochemistry associated with specific selection of aptamer for CAP. When the measurement of the fabricated nanomaterial chip was carried out using a three-electrode system, we found a highly specific and stable detection of chloramphenicol that ranged between 0.1 pM and 1 µM, with the detection limit of 0.1 pM. In addition, we obtained satisfactory results when real sample were used to validate the potential of photoelectrochemical (PEC) aptasensor for detecting chloramphenicol content in milk. Our results demonstrate that photoelectrochemical aptasensor is conducive to the development of less toxic multifunctional nanomaterials, making the biosensor more robust and environmentally friendly. Therefore, photoelectrochemical aptasensor can be widely applied in the field of environmental monitoring.


Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Nanotubos , Técnicas Electroquímicas , Límite de Detección , Titanio
18.
ACS Omega ; 7(2): 1819-1826, 2022 Jan 18.
Artículo en Inglés | MEDLINE | ID: mdl-35036821

RESUMEN

We report a novel design of chamber-based digital polymerase chain reaction (cdPCR) chip structure. Using a wet etching process and silicon-glass bonding, the chamber size can be adjusted independently of the process and more feasibly in a normal lab. In addition, the structure of the chip is optimized through hydrodynamic computer simulations to eliminate dead space when the sample is injected into the chip. The samples will be distributed to each separated microchambers for an isolated reaction based on Poisson distribution. Due to the difference in expansion coefficients, isolation of the sample in the microchambers by the oil phase on top ensures homogeneity and independence of the sample in the microchambers. The prepared microarray cdPCR chip enables high-throughput and high-sensitivity quantitative measurement of the SARS-CoV-2 virus gene and the mutant lung cancer gene. We applied the chip for the detection of different concentrations of the mix containing the open reading frame 1ab (ORF1ab) gene, the most specific and conservative gene region of the SARS-CoV-2 virus. In addition to this, we also successfully detected the fluorescence of the epidermal growth factor receptor (EGFR) mutant gene in independent microchambers. At a throughput of 46 200 microchambers, solution mixtures containing both genes were successfully tested quantitatively, with a detection limit of 10 copies/µL. Importantly, the chips are individually inexpensive and easy to industrialize. In addition, the microarray can provide a unified solution for other viral sequences, cancer marker assay development, and point-of-care testing (POCT).

19.
Plants (Basel) ; 11(24)2022 Dec 13.
Artículo en Inglés | MEDLINE | ID: mdl-36559607

RESUMEN

The 14-3-3 proteins, which are ubiquitous and highly conserved in eukaryotic cells, play an essential role in various areas of plant growth, development, and physiological processes. The tomato is one of the most valuable vegetable crops on the planet. The main objective of the present study was to perform genome-wide identification and analysis of the tomato 14-3-3 (SlTFT) family to investigate its response to different abiotic stresses and phytohormone treatments in order to provide valuable information for variety improvement. Here, 13 SlTFTs were identified using bioinformatics methods. Characterization showed that they were categorized into ε and non-ε groups with five and eight members, accounting for 38.5% and 61.5%, respectively. All the SlTFTs were hydrophilic, and most of them did not contain transmembrane structural domains. Meanwhile, the phylogeny of the SlTFTs had a strong correlation with the gene structure, conserved domains, and motifs. The SlTFTs showed non-random chromosomal distribution, and the promoter region contained more cis-acting elements related to abiotic stress tolerance and phytohormone responses. The results of the evolutionary analysis showed that the SlTFTs underwent negative purifying selection during evolution. Transcriptional profiling and gene expression pattern analysis showed that the expression levels of the SlTFTs varied considerably in different tissues and periods, and they played a specific role under various abiotic stresses and phytohormone treatments. Meanwhile, the constructed protein-based interaction network systematically broadens our understanding of SlTFTs. Finally, the virus-induced gene silencing of SlTFT4 affected the antioxidant and reactive oxygen species defense systems, increased the degree of cellular damage, and reduced salt resistance in tomatoes.

20.
Front Plant Sci ; 13: 1023696, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36570882

RESUMEN

As global soil salinization continues to intensify, there is a need to enhance salt tolerance in crops. Understanding the molecular mechanisms of tomato (Solanum lycopersicum) roots' adaptation to salt stress is of great significance to enhance its salt tolerance and promote its planting in saline soils. A combined analysis of the metabolome and transcriptome of S. lycopersicum roots under different periods of salt stress according to changes in phenotypic and root physiological indices revealed that different accumulated metabolites and differentially expressed genes (DEGs) associated with phenylpropanoid biosynthesis were significantly altered. The levels of phenylpropanoids increased and showed a dynamic trend with the duration of salt stress. Ferulic acid (FA) and spermidine (Spd) levels were substantially up-regulated at the initial and mid-late stages of salt stress, respectively, and were significantly correlated with the expression of the corresponding synthetic genes. The results of canonical correlation analysis screening of highly correlated DEGs and construction of regulatory relationship networks with transcription factors (TFs) for FA and Spd, respectively, showed that the obtained target genes were regulated by most of the TFs, and TFs such as MYB, Dof, BPC, GRAS, and AP2/ERF might contribute to the regulation of FA and Spd content levels. Ultimately, FA and Spd attenuated the harm caused by salt stress in S. lycopersicum, and they may be key regulators of its salt tolerance. These findings uncover the dynamics and possible molecular mechanisms of phenylpropanoids during different salt stress periods, providing a basis for future studies and crop improvement.

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