RESUMEN
Photodynamic therapy (PDT) is a significant noninvasive therapeutic modality, but it is often limited in its application due to the restricted tissue penetration depth caused by the wavelength limitations of the light source. Two-photon (TP) fluorescence techniques are capable of having an excitation wavelength in the NIR region by absorbing two NIR photons simultaneously, which offers the potential to achieve higher spatial resolution for deep tissue imaging. Thus, the adoption of TP fluorescence techniques affords several discernible benefits for photodynamic therapy. Organic TP dyes possess a high fluorescence quantum yield. However, the biocompatibility of organic TP dyes is poor, and the method of coating organic TP dyes with silica can effectively overcome the limitations. Herein, based on the TP silica nanoparticles, a functionalized intelligent biogenic missile TP-SiNPs-G4(TMPyP4)-dsDNA(DOX)-Aptamer (TGTDDA) was developed for effective TP bioimaging and synergistic targeted photodynamic therapy and chemotherapy in tumors. First, the Sgc8 aptamer was used to target the PTK7 receptor on the surface of tumor cells. Under two-photon light irradiation, the intelligent biogenic missile can be activated for TP fluorescence imaging to identify tumor cells and the photosensitizer assembled on the nanoparticle surface can be activated for photodynamic therapy. Additionally, this intelligent biogenic missile enables the controlled release of doxorubicin (DOX). The innovative strategy substantially enhances the targeted therapeutic effectiveness of cancer cells. The intelligent biogenic missile provides an effective method for the early detection and treatment of tumors, which has a good application prospect in the real-time high-sensitivity diagnosis and treatment of tumors.
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Imagen Óptica , Fotoquimioterapia , Fotones , Fármacos Fotosensibilizantes , Humanos , Animales , Fármacos Fotosensibilizantes/química , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/uso terapéutico , Ratones , Nanopartículas/química , Doxorrubicina/química , Doxorrubicina/farmacología , Doxorrubicina/uso terapéutico , Dióxido de Silicio/química , Aptámeros de Nucleótidos/química , Colorantes Fluorescentes/química , Neoplasias/tratamiento farmacológico , Neoplasias/diagnóstico por imagen , Antineoplásicos/química , Antineoplásicos/farmacología , Ratones Desnudos , Línea Celular Tumoral , Ratones Endogámicos BALB CRESUMEN
Persistent luminescent nanoparticles (PLNPs) are excellent luminescent materials, and near-infrared PLNPs are efficiently applied for biosensing and bioimaging due to their advantages of no excitation, excellent light stability and long afterglow. However, due to interference from the complex environment within organisms, single-mode imaging methods often face limitations in selectivity, sensitivity, and accuracy. Therefore, it is desirable to construct a dual-mode imaging probe strategy with higher specificity and sensitivity for bioimaging. Magnetic resonance imaging (MRI) has been widely used in the field of bioimaging due to its advantages of high resolution, non-radiation and non-invasiveness. Here, by combining near-infrared PLNPs and manganese dioxide (MnO2) nanosheets, a sensitive and convenient dual-mode "turn on" bioimaging nanoprobe ZGC@MnO2 has been developed for long afterglow imaging and MRI of endogenous hydrogen peroxide (H2O2) in the tumor microenvironment (TME). The monitoring of H2O2 has garnered significant attention due to its crucial role in human pathologies. For the dual-mode "turn on" bioimaging nanoprobe, the near-infrared PLNPs of quasi-spherical ZnGa2O4:Cr (ZGC) nanoparticles were synthesized as luminophores, and MnO2 nanosheets were utilized as a fluorescence quencher, carrier and H2O2 recognizer. H2O2 in the TME could reduce MnO2 nanosheets to Mn2+ for MRI, and ZGC nanoparticles were released for long afterglow imaging. Finally, the ZGC@MnO2 nanoprobe exhibited a rapid response, an excellent signal-to-noise ratio and a limit of detection of 3.67 nM for endogenous H2O2 in the TME. This dual-mode approach enhances the detection sensitivity for endogenous H2O2, thereby facilitating the research of endogenous H2O2-associated diseases and clinical diagnostics.
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Peróxido de Hidrógeno , Imagen por Resonancia Magnética , Compuestos de Manganeso , Nanopartículas , Óxidos , Microambiente Tumoral , Peróxido de Hidrógeno/química , Humanos , Imagen por Resonancia Magnética/métodos , Compuestos de Manganeso/química , Óxidos/química , Nanopartículas/química , Animales , Ratones , Células HeLa , Límite de DetecciónRESUMEN
The discovery of reliable biomarkers is essential for early diagnosis, treatment, and prognosis assessment of diseases. Many research studies have shown that circRNA is a potential biomarker for diagnosis and prognosis of diseases. However, in situ monitoring circRNA in live cells is still a challenge at present, which brings a major limitation to the development and verification of circRNA as a disease biomarker. In this study, a catalytic hairpin assembly (CHA) reaction-based DNA octahedral amplifier (DOA) was developed for fluorescence resonance energy transfer (FRET) detection and bioimaging of circRNA in living cells. The DOA was first produced by self-assembling a DNA octahedron with six customized single-stranded DNAs, and two hairpins H1 (Cy3) and H2 (Cy5) were then hybridized to four vertices of the DNA octahedron. Idiopathic pulmonary fibrosis (IPF)-related circHIPK3 was used as the target. Once the CHA reaction from H1 and H2 on DOA was activated by a sequence-specific back-splice junction (BSJ) of circHIPK3, a significant FRET signal can be obtained from Cy3 to Cy5. The circHIPK3 was subsequently released to cause the next CHA reaction. Because the DOA has the advantages of the spatial-confinement effect, resistance to nuclease degradation and easy penetration into cells, rapid and excellent signal amplification FRET detection and bioimaging of endogenous circHIPK3 can be achieved in various cells. This study provides a high-precision assay platform to explore the possibility of using circRNA as a biomarker, and it is valuable for circRNA-related early diagnosis and treatment of diseases.
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Técnicas Biosensibles , Carbocianinas , MicroARNs , MicroARNs/genética , ARN Circular/genética , ADN/genética , Biomarcadores , Técnicas Biosensibles/métodos , Límite de DetecciónRESUMEN
Hairy tofu is a famous Chinese snack that is made from soybeans and rich in various nutrients. In order to further explore the antioxidant peptides of hairy tofu hydrolysates, seven proteases were used to hydrolyze hairy tofu. The results of in vitro radical scavenging activity showed that hairy tofu hydrolysates obtained by pancreatin exhibited the highest antioxidant activity. After Sephadex G-25 gel filtration and reversed-phase high-performance liquid chromatography (RP-HPLC), 97 peptides were identified in the most antioxidant fraction using liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). Among them, nine peptides were synthesized and their antioxidant activities were assessed using a H2O2-induced oxidative 293T cell model. Finally, four peptides (QCESHK, LAWNEGR, NLQGENEWDQK, and FTEMWR) at concentrations of < 50 µg/ml significantly decreased the malondialdehyde content compared with the model group, displaying in vivo antioxidant activity and low cytotoxicity. Overall, this research provided the choice of using hairy tofu peptides as antioxidant products in the pharmaceutical and food industries.
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Antioxidantes , Péptidos , Humanos , Antioxidantes/química , Antioxidantes/farmacología , Cromatografía Líquida de Alta Presión , Células HEK293 , Peróxido de Hidrógeno , Hidrólisis , Péptidos/química , Péptidos/farmacología , Péptidos/aislamiento & purificación , Alimentos de Soja/análisisRESUMEN
Bioimaging is widely used in various fields of modern medicine. Fluorescence imaging has the advantages of high sensitivity, high selectivity, noninvasiveness, in situ imaging, and so on. However, one-photon (OP) fluorescence imaging has problems, such as low tissue penetration depth and low spatiotemporal resolution. These disadvantages can be solved by two-photon (TP) fluorescence imaging. However, TP imaging still uses fluorescence intensity as a signal. The complexity of organisms will inevitably affect the change of fluorescence intensity, cause false-positive signals, and affect the accuracy of the results obtained. Fluorescence lifetime imaging (FLIM) is different from other kinds of fluorescence imaging, which is an intrinsic property of the material and independent of the material concentration and fluorescence intensity. FLIM can effectively avoid the fluctuation of TP imaging based on fluorescence intensity and the interference of autofluorescence. Therefore, based on silica-coated gold nanoclusters (AuNCs@SiO2) combined with nucleic acid probes, the dual-mode nanoprobe platform was constructed for TP and FLIM imaging of intracellular endogenous miRNA-21 for the first time. First, the dual-mode nanoprobe used a dual fluorescence quencher of BHQ2 and graphene oxide (GO), which has a high signal-to-noise ratio and anti-interference. Second, the dual-mode nanoprobe can detect miR-21 with high sensitivity and selectivity in vitro, with a detection limit of 0.91 nM. Finally, the dual-mode nanoprobes performed satisfactory TP fluorescence imaging (330.0 µm penetration depth) and FLIM (τave = 50.0 ns) of endogenous miR-21 in living cells and tissues. The dual-mode platforms have promising applications in miRNA-based early detection and therapy and hold much promise for improving clinical efficacy.
RESUMEN
The rapid, simultaneous, sensitive detection of the targets has important application prospects for disease diagnosis and biomedical studies. However, in practical applications, the content of the targets is usually very low, and signal amplification strategies are often needed to improve the detection sensitivity. DNAzyme-driven DNA walkers are an excellent signal amplification strategy due to their outstanding specificity and sensitivity. Food-borne pathogens have always been a foremost threat to human health, and it is an urgent demand to develop a simple, rapid, sensitive, and portable detection method for food-borne pathogens. In addition, there are various species of pathogens, and it is difficult to simultaneously detect multiple pathogens by a single DNA walker. For this reason, a substrate strand with three rA cleavage sites was cleverly designed, and a multivalent DNA walker sensor combined with the microfluidic chip technology was proposed for the simultaneous, rapid, sensitive analysis of Vibrio parahaemolyticus, Salmonella typhimurium, and Staphylococcus aureus. The developed sensor could be used to detect pathogens simultaneously and efficiently with low detection limits and wide detection ranges. Moreover, the combination of gold stirring rod enrichment and DNA walker achieved double amplification, which greatly improved the detection sensitivity. More importantly, by changing the design of the substrate chain, the sensor was expected to be used to detect other targets, thus broadening the scope of practical applications. Therefore, the sensor can build novel detection tool platforms in the field of biosensing.
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Técnicas Biosensibles , Microfluídica , Humanos , Técnicas de Amplificación de Ácido Nucleico/métodos , Técnicas Biosensibles/métodos , ADN/genética , ADN/química , Análisis de Secuencia por Matrices de Oligonucleótidos , Límite de DetecciónRESUMEN
Rapid, simultaneous, and sensitive detection of biomolecules has important application prospects in disease diagnosis and biomedical research. However, because the content of intracellular endogenous target biomolecules is usually very low, traditional detection methods can't be used for effective detection and imaging, and to enhance the detection sensitivity, signal amplification strategies are frequently required. The hybridization chain reaction (HCR) has been used to detect many disease biomarkers because of its simple operation, good reproducibility, and no enzyme involvement. Although HCR signal amplification methods have been employed to detect and image intracellular biomolecules, there are still false positive signals. Therefore, a target-triggered enzyme-free amplification system (GHCR system) was developed, as a fluorescent AND-gated sensing platform for intracellular target probing. The false positive signals can be well avoided and the accuracy of detection and imaging can be improved by using the design of the AND gate. Two cancer markers, GSH and miR-1246, were used as two orthogonal inputs for the AND gated probe. The AND-gated probe only works when GSH and miR-1246 are the inputs at the same time, and FRET signals can be the output. In addition to the use of AND-gated imaging, FRET-based high-precision ratiometric fluorescence imaging was employed. FRET-based ratiometric fluorescent probes have a higher ability to resist interference from the intracellular environment, they can avoid false positive signals well, and they are expected to have good specificity. Due to the advantages of HCR, AND-gated, and FRET fluorescent probes, the GHCR system exhibited highly efficient AND-gated FRET bioimaging for intracellular endogenous miRNAs with a lower detection limit of 18 pM, which benefits the applications of ratiometric intracellular biosensing and bioimaging and offers a novel concept for advancing the diagnosis and therapeutic strategies in the field of cancer.
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Investigación Biomédica , MicroARNs , Neoplasias , Humanos , Colorantes Fluorescentes , Reproducibilidad de los Resultados , MicroARNs/genética , Neoplasias/diagnóstico por imagenRESUMEN
Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease, caused by severe fever with thrombocytopenia syndrome virus (SFTSV), which is primarily transmitted via tick bites. Clusters of SFTS caused by human-to-human transmission have been reported both at home and abroad, mainly focused on the transmission or exposure modes. However, the correlation between SFTS clusters and viral genotypes has not been investigated. This study mainly reported two clusters of SFTS in Xinyang City, Henan Province, from 2022 to 2023, discussed the possible route of person-to-person transmission of SFTSV infection and analyzed the association between SFTS clusters and virus genotypes. We found that two groups of SFTSV in two clusters were clustered separately into different genotypes through viral sequence analysis of 4 confirmed patients. We also performed phylogenetic analysis, after including SFTSV sequences obtained from SFTS clusters deposited in the GenBank. Three SFTSV genotypes have been reported among cases of human-to-human transmission, suggesting that the occurrence of SFTS clusters may not be related to SFTSV genotypes. This study provided genetic evidence for revealing the chain of human-to-human transmission of SFTS clusters, indicating that contact with patients' blood is an important transmission route of SFTSV. The findings laid the foundation for preventing and controlling human-to-human transmission of SFTS.
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Genotipo , Phlebovirus , Filogenia , Síndrome de Trombocitopenia Febril Grave , Humanos , Phlebovirus/genética , Síndrome de Trombocitopenia Febril Grave/virología , Síndrome de Trombocitopenia Febril Grave/transmisión , China/epidemiología , Masculino , FemeninoRESUMEN
Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tick-borne disease with a high case fatality rate. Few studies have been performed on bacterial or fungal coinfections or the effect of antibiotic therapy. A retrospective, observational study was performed to assess the prevalence of bacterial and fungal coinfections in patients hospitalized for SFTSV infection. The most commonly involved microorganisms and the effect of antimicrobial therapy were determined by the site and source of infection. A total of 1201 patients hospitalized with SFTSV infection were included; 359 (29.9%) had microbiologically confirmed infections, comprised of 292 with community-acquired infections (CAIs) and 67 with healthcare-associated infections (HAIs). Death was independently associated with HAIs, with a more significant effect than that observed for CAIs. For bacterial infections, only those acquired in hospitals were associated with fatal outcomes, while fungal infection, whether acquired in hospital or community, was related to an increased risk of fatal outcomes. The infections in the respiratory tract and bloodstream were associated with a higher risk of death than that in the urinary tract. Both antibiotic and antifungal treatments were associated with improved survival for CAIs, while for HAIs, only antibiotic therapy was related to improved survival, and no effect from antifungal therapy was observed. Early administration of glucocorticoids was associated with an increased risk of HAIs. The study provided novel clinical and epidemiological data and revealed risk factors, such as bacterial coinfections, fungal coinfections, infection sources, and treatment strategies associated with SFTS deaths/survival. This report might be helpful in curing SFTS and reducing fatal SFTS.
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Infecciones por Bunyaviridae , Coinfección , Phlebovirus , Síndrome de Trombocitopenia Febril Grave , Antibacterianos/uso terapéutico , Antifúngicos/uso terapéutico , Infecciones por Bunyaviridae/epidemiología , Coinfección/epidemiología , Humanos , Estudios RetrospectivosRESUMEN
Biological fluorescence imaging technologies have attracted a lot of attention and have been widely used in biomedical fields. Compared with other technologies, fluorescence imaging has a lower cost, higher sensitivity, and easier operation. However, due to the disadvantages of one-photon (OP) fluorescence imaging, such as low spatial and poor temporal resolution and poor tissue permeability depth, the application of OP fluorescence imaging has some limitations. Though two-photon (TP) fluorescence imaging can well overcome these shortcomings of OP, the single-mode imaging remains deficient. Therefore, dual-mode imaging combined with TP imaging and magnetic resonance imaging (MRI) can make up for the deficiency well, which make dual-mode imaging for the early diagnosis of diseases more accurate. Hence, a dual-mode nanoprobe TP-CQDs@MnO2 was designed for probing the fluorescence/MR dual-mode imaging strategy of intracellular H+ by using TP-CQDs (two photon-carbon quantum dots) and MnO2 nanosheets. The MnO2 nanosheets treated as fluorescence quenching agents of TP-CQDs exhibited a supersensitive response to H+, which made the fluorescence signals turn "off" to "on" for TP fluorescence imaging, in the meantime, large amounts of Mn2+ were generated for MRI. A dual-mode nanoprobe TP-CQDs@MnO2 can monitor intracellular wide pH (4.0-8.0), and the fluorescence intensity of TP-CQDs@MnO2 has recovered up to more than six times and the corresponding results of MRI were satisfactory. TP fluorescence imaging of cells and tissues showed higher detection sensitivity and deeper tissue penetration (240.0 µm) than OP. The dual-mode imaging platform hold great promise for pH-related early diagnosis and treatment, which has great potential to improve clinical efficacy.
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Compuestos de Manganeso , Puntos Cuánticos , Concentración de Iones de Hidrógeno , Imagen por Resonancia Magnética , Imagen Óptica , ÓxidosRESUMEN
Two-photon fluorescence imaging is one of the most attractive imaging techniques for monitoring important biomolecules in the biomedical field due to its advantages of low light scattering, high penetration depth, and suppressed photodamage/phototoxicity under near-infrared excitation. However, in actual biological imaging, organic two-photon fluorescent dyes have disadvantages such as high biological toxicity and their fluorescence efficiency is easily affected by the complex environment in organisms. In this study, a novel nanoprobe platform with two-photon dye-doped silica nanoparticles was developed for FRET-based ratiometric biosensing and bioimaging, with endogenous ATP chosen as the target for detection. The nanoprobe has three components: (1) a two-photon dye-doped silica nanoparticle core, which serves as an energy donor for FRET; (2) amino-modified hairpin primers with carboxy fluorescein as an energy acceptor for FRET; (3) an aptamer acting as a recognition unit to realize the probing function. The nanoprobe showed ratiometric fluorescence responses for ATP detection with high sensitivity and high selectivity in vivo. Moreover, the nanoprobe showed satisfactory ratiometric two-photon fluorescence imaging of endogenous ATP in living cells and tissues (penetration depth of 190 nm). These results indicated that novel two-photon silica nanoparticles can be constructed by doping a two-photon fluorescent dye into silica nanoparticles, and they can effectively solve the disadvantages of two-photon fluorescent dyes. These excellent performances indicate that this novel nanoprobe platform will become a very valuable molecular imaging tool, which can be widely used in the biomedical field for drug screening and disease diagnosis and other related research.
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Nanopartículas , Dióxido de Silicio , Adenosina Trifosfato , Transferencia Resonante de Energía de Fluorescencia , Colorantes Fluorescentes/toxicidad , Nanopartículas/toxicidad , Fotones , Dióxido de Silicio/toxicidadRESUMEN
The novel theranostic nanosystems based on two-photon fluorescence can achieve higher spatial resolution of deep tissue imaging for simultaneous diagnosis and therapy of a variety of cancers. Herein, we have designed and prepared FRET-based two-photon mesoporous silica nanoparticles (MTP-MSNs) for single-excitation multiplexed intracellular imaging and targeted cancer therapy for the first time. This nanosystem includes two constituents, containing (1) multicolor two-photon mesoporous silica nanoparticles and (2) cancer cell-targeting aptamers that act as gatekeepers for MTP-MSNs. After incubation with cancer cells, the Dox-loaded and aptamer-capped MTP-MSNs could be internalized into the cells, opening the pores and releasing the drug. Furthermore, using two-photon multicolor fluorescence, MTP-MSNs could serve as good contrast agents for multicolor two-photon intracellular imaging with increased imaging depth and improved spatial localization of tissue. In sum, these multicolor MTP-MSNs provide a promising system for traceable targeted cancer therapy with further applications in multiplex intracellular imaging and the screening of drug.
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Microscopía de Fluorescencia por Excitación Multifotónica/métodos , Nanopartículas/química , Neoplasias/diagnóstico , Animales , Aptámeros de Nucleótidos/química , Supervivencia Celular/efectos de los fármacos , Medios de Contraste/química , Doxorrubicina/química , Doxorrubicina/farmacología , Doxorrubicina/uso terapéutico , Portadores de Fármacos/química , Transferencia Resonante de Energía de Fluorescencia , Humanos , Rayos Láser , Hígado/efectos de los fármacos , Hígado/patología , Células MCF-7 , Neoplasias/tratamiento farmacológico , Oligodesoxirribonucleótidos/química , Porosidad , Ratas , Dióxido de Silicio/química , Nanomedicina TeranósticaRESUMEN
Sound conditioning (SC) is defined as "toughening" to lower levels of sound over time, which reduces a subsequent noise-induced threshold shift. Although the protective effect of SC in mammals is generally understood, the exact mechanisms involved have not yet been elucidated. To confirm the protective effect of SC against noise exposure (NE) and the stress-related signaling pathway of its rescue, we observed target molecule changes caused by SC of low frequency prior to NE as well as histology analysis in vivo and verified the suggested mechanisms in SGNs in vitro. Further, we investigated the potential role of Hsp70 and Bmi1 in SC by targeting SOD1 and SOD2 which are regulated by the FoxO1 signaling pathway based on mitochondrial function and reactive oxygen species (ROS) levels. Finally, we sought to identify the possible molecular mechanisms associated with the beneficial effects of SC against noise-induced trauma. Data from the rat model were evaluated by western blot, immunofluorescence, and RT-PCR. The results revealed that SC upregulated Hsp70, Bmi1, FoxO1, SOD1, and SOD2 expression in spiral ganglion neurons (SGNs). Moreover, the auditory brainstem responses (ABRs) and electron microscopy revealed that SC could protect against acute acoustic trauma (AAT) based on a significant reduction of hearing impairment and visible reduction in outer hair cell loss as well as ultrastructural changes in OHCs and SGNs. Collectively, these results suggested that the contribution of Bmi1 toward decreased sensitivity to noise-induced trauma following SC was triggered by Hsp70 induction and associated with enhancement of the antioxidant system and decreased mitochondrial superoxide accumulation. This contribution of Bmi1 was achieved by direct targeting of SOD1 and SOD2, which was regulated by FoxO1. Therefore, the Hsp70/Bmi1-FoxO1-SOD signaling pathway might contribute to the protective effect of SC against AAT in a rat model.
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Pérdida Auditiva Provocada por Ruido/metabolismo , Transducción de Señal , Estimulación Acústica , Animales , Potenciales Evocados Auditivos del Tronco Encefálico , Proteínas del Choque Térmico HSP72/metabolismo , Células Ciliadas Auditivas/ultraestructura , Pérdida Auditiva Provocada por Ruido/prevención & control , Masculino , Proteínas del Tejido Nervioso/metabolismo , Complejo Represivo Polycomb 1/metabolismo , Ratas Sprague-Dawley , Superóxido Dismutasa-1/metabolismoRESUMEN
The present study investigated the chemical composition, antioxidant, antimicrobial, and anti-inflammatory activities of essential oil (EO) derived from the wild rhizomes of Atractylodes macrocephala Koidz. (AMA) growing in Qimen County (eastern China). GC/MS analysis identified fifteen compounds, representing 92.55 % of AMA EO. The major compounds were atractylone (39.22 %), ß-eudesmol (27.70 %), thymol (5.74 %), hinesol (5.50 %), and 11-isopropylidenetricyclo[4.3.1.1(2,5)]undec-3-en-10-one (4.71 %). Ferricyanide reducing, 1,1-diphenyl-2-picyrlhydrazyl (DPPH) and 3-ethyl-benzothiazoline-6-sulfonic acid (ABTS) scavenging assays revealed that AMA EO exhibited strong antioxidant capacities. Additionally, AMA EO showed inhibitory effects on growth of Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, and Bacillus subtilis, with the minimum inhibitory concentrations (MIC) ranging from 0.5 to 2.0â mg/mL. Treatments with AMA EO also significantly inhibited nitric oxide (NO) and prostaglandin E2 (PGE2 ) production in lipopolysaccharide-stimulated RAW264.7 cells, indicating anti-inflammatory activity of AMA EO. Furthermore, treatments with AMA EO decreased the transcriptional levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), which might be the molecular mechanisms underlying its anti-inflammatory effects. Overall, these results provide a theoretical basis for further study and application of AMA EO in food and medicine products.
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Antibacterianos/farmacología , Antiinflamatorios/farmacología , Antioxidantes/farmacología , Atractylodes/química , Aceites Volátiles/farmacología , Rizoma/química , Animales , Cromatografía de Gases y Espectrometría de Masas , Ratones , Pruebas de Sensibilidad Microbiana , Células RAW 264.7RESUMEN
An enzyme-free fluorometric assay is described that accomplishes dual signal amplification by making use of a two stirring bars. Two Y-shaped DNA probes were designed and placed on the bars. When the target (with chloramphenicol as model analyte) is added, it triggers target recycling and simultaneously catalyzes hairpin assembly (CHA). A large fraction of DNA primers is released by the analyte from the bar to the supernatant and open hairpins with G-quadruplex DNA sequence. The G-quadruplex can specifically bind thioflavin T (ThT) to emit fluorescence (with excitation/emission maxima at 445 and 485 nm) for quantification of chloramphenicol. An enzyme is not needed. ThT is added to the system as a fluorescent DNA probe. All this strongly reduces the cost for sensor construction and usage. The dual signal amplification steps occur simultaneously which reduces the detection time. The assay was successfully employed to the determination of CAP in spiked milk and fish samples within 60 min and with a 16 pM limit of detection (at S/N = 3). Graphical abstract Schematic representation of a new method for the detection of chloramphenicol by using two stirring bars. It is based on target recycling and catalyzed hairpin assembly amplification. CAP: chloramphenicol, ThT: thioflavin T, CHA: catalyzed hairpin assembly.
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Cloranfenicol/análisis , Fluorometría/métodos , Sondas de ADN , Colorantes Fluorescentes , Fluorometría/instrumentación , G-Cuádruplex , Límite de DetecciónRESUMEN
A simple and highly sensitive fluorometric method is described for the determination of the antibiotic kanamycin (Kana) in food. Dual signal amplification is accomplished by making use of double Y-shaped aptamer DNA probes acting as a capture probes and signal amplification probes. The DNA probes were immobilized on a gold bar and on a magnetic bar, respectively. On addition of Kana, the Y-shaped aptamer probe captures Kana and then is disassembled to release two single-stranded DNAs. These trigger target recycling and HCR between the two bars simultaneously. As a result, many long duplex DNA chains are formed in the supernatant. After pulling out the bars and adding the fluorescent intercalating probe SYBR Green I, strong fluorescence (with excitation/emission peaks at 497/525 nm) is induced. The use of such double Y-shaped DNA probes obviously overcomes the unspecific signal amplification by HCR which increases selectivity and sensitivity. This is due to the fact that the hairpin of HCR is separated in being present in different arms of the Y-shaped probe. Under the optimal conditions, the assay has a limit of 0.45 pg·mL-1 for Kana. It was applied to analyze spiked milk, fish and pork samples. Graphical abstract The scheme represents a sensitive fluorometric aptamer-based method to detect kanamycin (Kana). It is making use of a double stirring bar-assisted dual amplification strategy with zero background. Abbreviations: apt: aptamer, AuNPs: gold nanoparticles, HCR: hybridization chain reaction.
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Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Sondas de ADN/química , Fluorometría/métodos , Oro/química , Kanamicina/análisis , Nanopartículas de Magnetita/química , Análisis de los Alimentos , Contaminación de Alimentos/análisis , Kanamicina/químicaRESUMEN
Recently, it has been crucial to be able to detect and quantify small molecular targets simultaneously in biological samples. Herein, a simple and conventional double-T type microchip electrophoresis (MCE) based platform for the multiplex detection of quality indicator molecule targets in urine, using ampicillin (AMPI), adenosine triphosphate (ATP) and estradiol (E2) as models, was developed. Several programmable hairpin probes (PHPs) were designed for detecting different targets and triggering isothermal polymerase-catalyzed target recycling (IPCTR) for signal amplification. Based on the target-responsive aptamer structure of PHP (Domain I), target recognition can induce PHP conformational transition and produce extension duplex DNA (dsDNA), assisted by primers & Bst polymerase. Afterwards, the target can be displaced to react with another PHP and initiate the next cycle. After several rounds of reaction, the dsDNA can be produced in large amounts by IPCTR. Three targets can be simultaneously converted to dsDNA fragments with different lengths, which can be separated and detected using MCE. Thus, a simple double-T type MCE based platform was successfully built for the homogeneous detection of multiplex targets in one channel. Under optimal conditions, the assay exhibited high throughput (48 samples per hour at most, not including reaction time) and sensitivity to three targets in urine with a detection limit of 1 nM (ATP), 0.05 nM (AMPI) and 0.1 nM (E2) respectively. The multiplex assay was successfully employed for the above three targets in several urine samples and combined the advantages of the high specificity of programmable hairpin probes, the excellent signal amplification of IPCTR, and the high through-put of MCE which can be employed for screening in biochemical analysis.
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Aptámeros de Nucleótidos , Electroforesis por Microchip , Urinálisis/métodos , Adenosina Trifosfato/orina , Ampicilina/orina , Estradiol/orina , Humanos , Límite de DetecciónRESUMEN
BACKGROUND: Group A streptococcus (GAS) infections and poststreptococcal sequelae remain a health problem worldwide, which necessitates searching for an effective vaccine, while no licensed GAS vaccine is available. We have developed a divalent peptide vaccine composed of 84 amino acids to cover the main GAS serotypes (M1 and M12 streptococci) in China, and herein, we aimed to evaluate immunogenicity and safety of this vaccine. METHODS: Mice were immunized with the vaccine. ELISA, indirect bactericidal test, and immunofluorescent assay were used to study immunogenicity. GAS challenge assay was used to test the protective effect. Safety was tested by histopathological analysis. RESULTS: Immunized group mice (n=16) developed higher titer antibody after immunization than nonimmunized group mice (n=16) did. This antibody can deposit on the surface of GAS and promote killing of GAS, resulting in 93.1% decrease of M1 GAS and 89.5% of M12 GAS. When challenged with M1 and M12 streptococci, immunized group mice had a higher survival rate (87.5% and 75%) than nonimmunized group mice (37.5% and 25%). No autoimmune reactions were detected on organs of mice. CONCLUSION: The results suggest that this vaccine shows fair immunogenicity and safety, which will lead our research on GAS vaccine into clinical trial.
RESUMEN
Ascorbic acid (AA), as one of the most important vitamins, participates in various physiological reactions in the human body and is implicated with many diseases. Therefore, the development of effective methods for monitoring the AA level in living systems is of great significance. Up to date, various technologies have been developed for the detection of AA. However, few methods can realize the direct detection of endogenous AA in living cells. In this work, we for the first time reported that near-infrared (NIR) graphene quantum dots (GQD) possessed good two-photon fluorescence properties with a NIR emission at 660 nm upon exciting with 810 nm femtosecond pulses and a two-photon (TP) excitation action cross-section (δΦ) of 25.12 GM. They were then employed to construct a TP nanoprobe for detection and bioimaging of endogenous AA in living cells. In this nanosystem, NIR GQDs (NGs), which exhibited lower fluorescence background in living system to afford improved fluorescence imaging resolution, were acted as fluorescence reporters. Also CoOOH nanoflakes were chosen as fluorescence quenchers by forming on the surface of NGs. Once AA was introduced, CoOOH was reduced to Co2+, which resulted in a "turn-on" fluorescence signal of NGs. The proposed nanoprobe demonstrated high sensitivity toward AA, with the observed LOD of 270 nM. It also showed high selectivity to AA with excellent photostability. Moreover, the nanoprobe was successfully used for TP imaging of endogenous AA in living cells as well as deep tissue imaging.
Asunto(s)
Ácido Ascórbico/análisis , Colorantes Fluorescentes/química , Grafito/química , Nanopartículas/química , Imagen Óptica , Fotones , Puntos Cuánticos/química , Células HeLa , Humanos , Tamaño de la Partícula , Propiedades de SuperficieRESUMEN
Semiconductor quantum dots have attracted extensive interest in the biosensing area because of their properties, such as narrow and symmetric emission with tunable colors, high quantum yield, high stability and controllable morphology. The introduction of various reactive functional groups on the surface of semiconductor quantum dots allows one to conjugate a spectrum of ligands, antibodies, peptides, or nucleic acids for broader and smarter applications. Among these ligands, aptamers exhibit many advantages including small size, high chemical stability, simple synthesis with high batch-to-batch consistency and convenient modification. More importantly, it is easy to introduce nucleic acid amplification strategies and/or nanomaterials to improve the sensitivity of aptamer-based sensing systems. Therefore, the combination of semiconductor quantum dots and aptamers brings more opportunities in bioanalysis. Here we summarize recent advances on aptamer-functionalized semiconductor quantum dots in biosensing applications. Firstly, we discuss the properties and structure of semiconductor quantum dots and aptamers. Then, the applications of biosensors based on aptamer-modified semiconductor quantum dots by different signal transducing mechanisms, including optical, electrochemical and electrogenerated chemiluminescence approaches, is discussed. Finally, our perspectives on the challenges and opportunities in this promising field are provided.