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Developing highly sensitive and selective methods that incorporate specific recognition elements is crucial for detecting small molecules because of the limited availability of small molecule antibodies and the challenges in obtaining sensitive signals. In this study, a generalizable photoelectrochemical-colorimetric dual-mode sensing platform was constructed based on the synergistic effects of a molecularly imprinted polymer (MIP)-aptamer sandwich structure and nanoenzymes. The MIP functionalized peroxidase-like Fe3O4 (Fe3O4@MIPs) and alkaline phosphatase mimic Zr-MOF labeled aptamer (Zr-mof@Apt) were used as the recognition elements. By selectively accumulating dibutyl phthalate (DBP), a small molecule target model, on Fe3O4@MIPs, the formation of Zr-MOF@Apt-DBP- Fe3O4@MIPs sandwich structure was triggered. Fe3O4@MIPs oxidized TMB to form blue-colored oxTMB. However, upon selective accumulation of DBP, the catalytic activity of Fe3O4@MIPs was inhibited, resulting in a lighter color that was detectable by the colorimetric method. Additionally, Zr-mof@Apt effectively catalyzed the hydrolysis of L-Ascorbic acid 2-phosphate sesquimagnesium salt hydrate (AAPS), generating ascorbic acid (AA) that could neutralize the photogenerated holes to decrease the photocurrent signals for PEC sensing and reduce oxTMB for colorimetric testing. The dual-mode platform showed strong linearity for different concentrations of DBP from 1.0 pM to 10 µM (PEC) and 0.1 nM to 0.5 µM (colorimetry). The detection limits were 0.263 nM (PEC) and 30.1 nM (colorimetry) (S/N = 3), respectively. The integration of dual-signal measurement mode and sandwich recognition strategy provided a sensitive and accurate platform for the detection of small molecules.
Asunto(s)
Técnicas Biosensibles , Polímeros Impresos Molecularmente , Colorimetría/métodos , Peroxidasa/química , PeroxidasasRESUMEN
In spite of organic-inorganic perovskite emerging as a novel efficient light-harvesting material owing to their superior optical properties, excitonic properties, and electrical conductivity, the related applications are severely limited for their poor stability and selectivity. Herein, we introduced hollow carbon spheres (HCSs) and 2-(perfluorohexyl) ethyl methacrylate (PFEM) based molecularly imprinted polymers (MIPs) to dual-functionalize CH3NH3PbI3. HCSs can provide perovskite load conditions, passivate perovskite defects, increase carrier transport and effectively improve its hydrophobicity. The perfluorinated organic compound based MIPs film can not only enhance the water and oxygen stability of perovskite, but also endow it specific selectivity. Moreover, it can reduce the photoexcited electron-hole pair recombination and prolong the electron lifetime. Benefiting from the synergistic sensitization of HCSs and MIPs, an ultrasensitive photoelectrochemical platform (MIPs@CH3NH3PbI3@HCSs/ITO) for cholesterol sensing was acquired with a very wide linear range of 5.0 × 10-14-5.0 × 10-8 mol/L and an extremely low detection limit of 2.39 × 10-15 mol/L. The designed PEC sensor exhibited good selectivity and stability, as well as practicality for real sample analysis. The present work extended the development of the high-performance perovskite and showed its broad application prospect for advanced PEC construction.
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Técnicas Biosensibles , Impresión Molecular , Impresión Molecular/métodos , Técnicas Electroquímicas/métodos , Técnicas Biosensibles/métodos , Carbono/químicaRESUMEN
A highly sensitive and selective split-type perovskite-based photoelectrochemical (PEC) platform was developed for measuring alkaline phosphatase (ALP) activity in milk and serum samples. ALP in the test sample hydrolyzed 2-phosphate sesquimagnesium salt hydrate (AAPS) in a 96-microwell plate to produce ascorbic acid (AA), a PEC electron donor. The resulting AA, which could preferentially annihilate the photogenerated holes, indirectly reflects ALP activity. The PEC used a cetyltrimethylammonium bromide (CTAB)-functionalized CH3NH3PbI3 (CTAB@CH3NH3PbI3) film as the cathode to monitor the controlled AA production. Due to the excellent photoelectric characteristics of the CH3NH3PbI3 perovskite and the split-type assay, excellent sensitivity and selectivity for ALP detection were obtained. Under the optimum experimental conditions, ALP activity with a limit of detection (LOD) of 2.6 × 10-4 U/L in a linear dynamic range of 10-3 â¼ 102 U/L was obtained. With its sensitive, rapid, and high-throughput detection capabilities, this split-type and label-free PEC platform has great potential for use in food and biomedical analysis.
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Fosfatasa Alcalina , Técnicas Biosensibles , Cetrimonio , Titanio , Electrodos , Límite de DetecciónRESUMEN
In this study, ß-Bi2O3 nanosheets functionalized with bisphenol A (BPA) synthetic receptors were developed by a simple molecular imprinting technology and applied as the photoelectric active material for the construction of a BPA photoelectrochemical (PEC) sensor. BPA was anchored on the surface of ß-Bi2O3 nanosheets via the self-polymerization of dopamine monomer in the presence of a BPA template. After the elution of BPA, the BPA molecular imprinted polymer (BPA synthetic receptors)-functionalized ß-Bi2O3 nanosheets (MIP/ß-Bi2O3) were obtained. Scanning electron microscopy (SEM) of MIP/ß-Bi2O3 revealed that the surface of ß-Bi2O3 nanosheets was covered with spherical particles, indicating the successful polymerization of the BPA imprinted layer. Under the best experimental conditions, the PEC sensor response was linearly proportional to the logarithm of BPA concentration in the range of 1.0 nM to 1.0 µM, and the detection limit was 0.179 nM. The method had high stability and good repeatability, and could be applied to the determination of BPA in standard water samples.
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The extension application of perovskites in aqueous media such as bioassays requires the development of a water-stable perovskite with a simple preparation process and low cost. However, the degradation of perovskites in aqueous solution is still a thorny problem. Here, we develop a methylimidazole-assisted two-step synthesis protocol to prepare CsPbBr3@PbBrOH nanorods with superior water stability and remarkable optical properties at room temperature. The synergy of 2-methylimidazole (2-MIM), an N-donor ligand, with water can not only facilitate CsPbBr3 formation and suppress CsPb2Br5 or Cs4PbBr6 formation, but also promote the formation of a PbBrOH shell capping CsPbBr3. 2-MIM is ionized into 2-MIM- in DMF and 2-MIM+ in water. They passivated the surface defects and changed the crystallization environment, leading to water-stable CsPbBr3@PbBrOH. The obtained CsPbBr3@PbBrOH nanorods can still maintain 91% PL intensity after being stored in water for more than 2 months. Furthermore, the CsPbBr3@PbBrOH nanorods show excellent stability in polar solvents, water, and phosphate buffer solution in a wide pH range, as well as better thermal and irradiation stability. In addition, the CsPbBr3@PbBrOH nanorods are further functionalized with polydopamine (PDA) for biomolecular immobilization and immunoassay studies. The resulting assay shows a detection limit of 0.003 ng mL-1 for IgG detection, illustrating important progress towards expanding fluorescence labeling application of perovskite nanomaterials for immunoassays.
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Nanoestructuras , Nanotubos , Óxidos , AguaRESUMEN
Long-term and excessive exposure to bisphenol A (BPA) has an extremely detrimental effect on human health and ecological system. Hence, there is an urgent need to develop a sensitive and selective sensor for precisely monitoring BPA levels. In this work, a flexible and tailor-made electrochemical sensor for BPA has been fabricated based on in situ photopolymerization of molecular imprinting on cadmium sulfide (CdS) modified carbon felt (MIP@CdS-CF). It is worth nothing that CdS acts as a photocatalyst to enhance the capacity of photopolymerization, accordingly upgrading imprinting efficiency. Meanwhile, carbon felt (CF) exhibits attractive merits in term of superior electrical conductivity, enlarged electrochemical active areas and unique flexibility. Consequently, the novel MIP@CdS-CF electrochemical sensor shows superior sensitivity, high selectivity, extraordinary reproducibility and stability to detect BPA. The detection limit is 1.5 nM, which is lower than those of previously reported electrochemical sensors for the detection of BPA. More importantly, this newly developed electrochemical sensor can be utilized for detecting BPA in plastic bottles with satisfactory results.
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Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) is an efficient tool for establishing genetic models including cellular models, and has facilitated unprecedented advancements in biomedical research. In both patients and cancer animal models, immune cells infiltrate the tumor microenvironment and some of them migrate to draining lymph nodes to exert antitumor effects. Among these immune cells, phagocytes such as macrophages and dendritic cells engulf tumor antigens prior to their crosstalk with T cells and elicit adaptive immune response against tumors. Melanoma cells are frequently used as a tumor model because of their relatively high level of somatic mutations and antigenicity. However, few genetic models have been developed using melanoma cell lines to track tumor cell phagocytosis, which is essential for understanding protective immune response in vivo. In this study, we used CRISPR/Cas9-mediated DNA cleavage and homologous recombination to develop a novel knock-in tool which expresses the ultra-bright fluorescent probe ZsGreen in YUMM1.7 melanoma cells. Using this novel tool, we measured the macrophagic engulfment of melanoma cells inside the tumor microenvironment. We also found that in tumor-grafted mice, a subset of dendritic cells efficiently engulfed YUMM1.7 cells and was preferentially trafficking tumor antigens to draining lymph nodes. In addition, we used this knock-in tool to assess the impact of a point mutation of CD11b on phagocytosis in the tumor microenvironment. Our results demonstrate that the ZsGreen-expressing YUMM1.7 melanoma model provides a valuable tool for the study of phagocytosis in vivo.
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Antígeno CD11b , Melanoma , Fagocitosis , Animales , Antígenos de Neoplasias , Antígeno CD11b/genética , Línea Celular , Línea Celular Tumoral , Colorantes Fluorescentes , Melanoma/genética , Ratones , Mutación Puntual , Microambiente TumoralRESUMEN
Here, a label-free perovskite-based photoelectrochemical (PEC) aptasensor was rationally designed for the displacement assay of dibutyl phthalate (DBP), a well-known endocrine disruptor, with the aid of cetyltrimethylammonium bromide (CTAB). In this method, CTAB significantly enhanced the PEC response and humidity resistance of the CH3NH3PbI3 perovskite by forming a protecting layer and passivating the X- and A-sites vacancies of CH3NH3PbI3. In addition, CTAB facilitated the immobilization of an aptamer through van der Waals and hydrophobicity forces, as well as the electrostatic interactions between the phosphate group of the aptamer and the cationic group of CTAB. When exposed to DBP in the affinity solution, the DBP aptamer was released from the electrode because the affinity between DBP and its aptamer competes with the interaction of the aptamer and CTAB. The displacement of the aptamer from the perovskite surface relieves the block effect and thus enhances the photoelectric signal of perovskite. By virtue of the good photoelectrochemical characters of CH3NH3PbI3 and the specific recognition ability of aptamer, the linear range of the PEC sensor was 1.0 × 10-13 to 1.0 × 10-8 M and the detection and quantification limits were down to 2.5 × 10-14 and 8.2 × 10-14 M (S/N = 3), respectively. This work offers a novel strategy for designing aptasensors for the detection of various targets and exhibits the marvelous potential of organic-inorganic perovskite in the field of PEC analysis.
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Aptámeros de Nucleótidos , Técnicas Biosensibles , Compuestos de Calcio , Cetrimonio , Dibutil Ftalato , Técnicas Electroquímicas/métodos , Límite de Detección , Óxidos , TitanioRESUMEN
This work designed a novel dioctyl phthalate (DOP) photoelectrochemical (PEC) sensor based on a molecularly imprinted polymer (MIP) modified Cu3(BTC)2@Cu2O heterostructure. In this work, a metal organic framework (MOF), Cu3(BTC)2, was coated on Cu2O through a simple immersion method to form a Cu3(BTC)2@Cu2O heterostructure. The heterostructure exhibited strong light adsorption ability, good stability and enhanced photocurrent under visible light irradiation. Using the Cu3(BTC)2@Cu2O heterostructure as the photoelectric converter, a PEC sensor was constructed by imprinting DOP on the heterostructure. Under the optimal experimental conditions, the PEC sensor showed a wide linear range from 25.0 pM-0.1 µM and a low detection limit of 9.15 pM. This method with good sensitivity, stability, selectivity and reproducibility in actual sample analyses showed promising applications of the MOF-based heterostructure in photoelectrochemical analysis fields.
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The aim of the present study was to investigate whether sulforaphane (SFN) and myricetin (Myr) synergistically induce apoptosis in adipocytes. The viability of mature 3T3L1 adipocytes treated with 40 µM SFN and/or 100 µM Myr was assessed using an MTT assay. Apoptosis was assessed by Hoechst 33258 nuclear staining, and by detection of singlestranded DNA using an enzymelinked immunosorbent assay. Compared with the effects of each compound alone, the combination of SFN and Myr synergistically reduced cell viability, induced apoptosis, increased proapoptotic Bcl2 associated X protein expression, decreased antiapoptotic Bcell lymphoma2 expression, enhanced Bcl2associated death promoter (Bad) translocation from the cytoplasm to the mitochondria, and reduced Bad phosphorylation at Ser112. These effects were accompanied by increased cleavage of caspase 3 and polyADPribosepolymerase. In addition, combined SFN and Myr treatment significantly decreased the protein expression levels of phosphorylated AKT serine/threonine kinase 1 (Akt) at Ser473, as well as the phosphorylation of the downstream protein ribosomal protein, S6 kinase ß1. Therefore, SFN plus Myr was a more potent inducer of apoptosis in 3T3L1 adipocytes than either compound alone. The results of the present study suggest that the mechanism of SNF/Myrinduced apoptosis involved activation of the Aktmediated mitochondrial apoptotic pathway. This may aid treatment of animal models of obesity and preclinical testing.
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Adipocitos/efectos de los fármacos , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Flavonoides/farmacología , Isotiocianatos/farmacología , Células 3T3-L1 , Adipocitos/citología , Adipocitos/metabolismo , Animales , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacos , SulfóxidosRESUMEN
Sulforaphane (SFN), an isothiocyanate isolated from cruciferous vegetables, possesses anti-oxidant and anti-cancer bioactivities. Moreover, SFN exerts its pro-apoptotic effects in some cancer lines. However, the effects and mechanisms of SFN on the regulation of apoptosis of adipocytes are still unknown. In this study, we found that SFN induced significant apoptosis in 3T3-L1 adipocytes and markedly decreased the cellular lipid content. Western blot demonstrated that SFN-induced apoptosis was mediated via the mitochondrial apoptosis pathway based on increased cleavage of poly-ADP-ribose-polymerase (PARP), release of cytochrome c into the cytoplasm, and activation of caspase-3, as well as decreased Bcl-2/Bax ratio. In addition, SFN markedly decreased phosphorylation of Akt and downstream proteins, p70s6k1 and Bad, and increased phosphorylation of ERK. Therefore, our findings clarified that SFN could induce 3T3-L1 adipocyte apoptosis via down-regulation of the Akt/p70s6k1/Bad pathway and up-regulation of the ERK pathway, suggesting SFN may be a promising agent for the treatment or prevention of obesity.
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Adipocitos/efectos de los fármacos , Fármacos Antiobesidad/farmacología , Apoptosis/efectos de los fármacos , Isotiocianatos/farmacología , Células 3T3-L1 , Adipocitos/citología , Adipocitos/metabolismo , Animales , Metabolismo de los Lípidos/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Modelos Biológicos , Fitoquímicos/farmacología , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Quinasas S6 Ribosómicas 70-kDa/antagonistas & inhibidores , Sulfóxidos , Proteína Letal Asociada a bcl/antagonistas & inhibidoresRESUMEN
The present investigation was undertaken to gain insight into the molecular mechanism by which myricetin induces apoptosis in human hepatocarcinoma HepG2 cells. Myricetin caused the disruption of mitochondrial membrane potential in a dose-dependent manner. Moreover, myricetin triggered translocation of the pro-apoptotic protein Bax to the mitochondria, downregulation of anti-apoptotic Bcl-2 expression and upregulated the expression of pro-apoptotic protein Bad in the mitochondria. The present study also showed that myricetin promoted the release of cytochrome C from mitochondria into the cytosol followed by an increase in the proteolytic activation of caspase-3 and the concomitant degradation of PARP protein. Additionally, western blot analysis showed that the Akt/p70s6k1 pathway was inhibited in myricetin-treated HepG2 cells, accordingly the phosphorylation of Bad at Ser136 was downregulated. Collectively, these findings indicate that myricetin induced apoptosis in HepG2 cell through mitochondria apoptotic pathway and Akt/p70s6k1/Bad signaling. Present results provide new information on the possible mechanisms for the anti-cancer activity of myricetin.
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Antineoplásicos Fitogénicos/farmacología , Flavonoides/farmacología , Regulación Neoplásica de la Expresión Génica , Mitocondrias/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Quinasas S6 Ribosómicas 70-kDa/genética , Proteína Letal Asociada a bcl/genética , Apoptosis/efectos de los fármacos , Caspasa 3/genética , Caspasa 3/metabolismo , Citocromos c/metabolismo , Relación Dosis-Respuesta a Droga , Células Hep G2 , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/metabolismo , Poli(ADP-Ribosa) Polimerasas/genética , Poli(ADP-Ribosa) Polimerasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Transducción de Señal , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismo , Proteína Letal Asociada a bcl/metabolismoRESUMEN
Myricetin, a naturally occurring flavonol, has been shown to inhibit the proliferation of human hepatoma HepG2 cells and to induce G2/M phase arrest. However, the underlying mechanisms of Myricetin activity have yet to be revealed. The aim of the present study was to clarify the molecular mechanisms of cell cycle arrest induced by myricetin in HepG2 cells. The MTT assay confirmed that exposure of HepG2 cells to myricetin triggered G2/M phase arrest. Western blot analysis showed that myricetin increased the protein levels of the p53/p21 cascade, and markedly decreased Cdc2 and cyclin B1 protein levels in HepG2 cells. Additionally, myricetin treatment resulted in the up-regulation of Thr14/Tyr15 phosphorylated (inactive) Cdc2 and p27, and the down-regulation of CDK7 kinase protein, as well as CDK7-mediated Thr161 phosphorylated (active) Cdc2. These data indicate that a decrease in cyclin B/Cdc2 complex activity mediated G2/M phase arrest induced by myricetin in HepG2 cells. This novel finding provides insight into the potential applications of myricetin in the treatment of hepatocellular carcinoma.