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1.
J Immunol ; 2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38847616

RESUMO

The cyclic GMP-AMP synthase (cGAS)-stimulator of IFN genes (STING) pathway is instrumental to antitumor immunity, yet the underlying molecular and cellular mechanisms are complex and still unfolding. A new paradigm suggests that cancer cells' cGAS-synthesized cGAMP can be transferred to tumor-infiltrating immune cells, eliciting STING-dependent IFN-ß response for antitumor immunity. Nevertheless, how the tumor microenvironment may shape this process remains unclear. In this study, we found that extracellular ATP, an immune regulatory molecule widely present in the tumor microenvironment, can potentiate cGAMP transfer, thereby boosting the STING signaling and IFN-ß response in murine macrophages and fibroblasts. Notably, genetic ablation or chemical inhibition of murine volume-regulation anion channel LRRC8/volume-regulated anion channel (VRAC), a recently identified cGAMP transporter, abolished ATP-potentiated cGAMP transfer and STING-dependent IFN-ß response, revealing a crucial role of LRRC8/VRAC in the cross-talk of extracellular ATP and cGAMP. Mechanistically, ATP activation of the P2X family receptors triggered Ca2+ influx and K+ efflux, promoting reactive oxygen species production. Moreover, ATP-evoked K+ efflux alleviated the phosphorylation of VRAC's obligate subunit LRRC8A/SWELL1 on S174. Mutagenesis studies indicated that the phosphorylation of S174 on LRRC8A could act as a checkpoint for VRAC in the steady state and a rheostat of ATP responsiveness. In an MC38-transplanted tumor model, systemically blocking CD39 and ENPP1, hydroxylases of extracellular ATP and cGAMP, respectively, elevated antitumor NK, NKT, and CD8+ T cell responses and restrained tumor growth in mice. Altogether, this study establishes a crucial role of ATP in facilitating LRRC8/VRAC transport cGAMP in the tumor microenvironment and provides new insight into harnessing cGAMP transfer for antitumor immunity.

2.
Anal Chem ; 96(24): 10092-10101, 2024 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-38833634

RESUMO

Tumor patients-derived organoids, as a promising preclinical prediction model, have been utilized to evaluate ex vivo drug responses for formulating optimal therapeutic strategies. Detecting adenosine triphosphate (ATP) has been widely used in existing organoid-based drug response tests. However, all commercial ATP detection kits containing the cell lysis procedure can only be applied for single time point ATP detection, resulting in the neglect of dynamic ATP variations in living cells. Meanwhile, due to the limited number of viable organoids from a single patient, it is impractical to exhaustively test all potential time points in search of optimal ones. In this work, a multifunctional microfluidic chip was developed to perform all procedures of organoid-based drug response tests, including establishment, culturing, drug treatment, and ATP monitoring of organoids. An ATP sensor was developed to facilitate the first successful attempt on whole-course monitoring the growth status of fragile organoids. To realize a clinically applicable automatic system for the drug testing of lung cancer, a microfluidic chip based automated system was developed to perform entire organoid-based drug response test, bridging the gap between laboratorial manipulation and clinical practices, as it outperformed previous methods by improving data repeatability, eliminating human error/sample loss, and more importantly, providing a more accurate and comprehensive evaluation of drug effects.


Assuntos
Trifosfato de Adenosina , Dispositivos Lab-On-A-Chip , Organoides , Humanos , Organoides/citologia , Organoides/efeitos dos fármacos , Organoides/metabolismo , Trifosfato de Adenosina/análise , Trifosfato de Adenosina/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Antineoplásicos/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Técnicas Analíticas Microfluídicas/instrumentação , Automação
3.
Cancer Immunol Immunother ; 73(6): 111, 2024 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-38668781

RESUMO

The increase in the detection rate of synchronous multiple primary lung cancer (MPLC) has posed remarkable clinical challenges due to the limited understanding of its pathogenesis and molecular features. Here, comprehensive comparisons of genomic and immunologic features between MPLC and solitary lung cancer nodule (SN), as well as different lesions of the same patient, were performed. Compared with SN, MPLC displayed a lower rate of EGFR mutation but higher rates of BRAF, MAP2K1, and MTOR mutation, which function exactly in the upstream and downstream of the same signaling pathway. Considerable heterogeneity in T cell receptor (TCR) repertoire exists among not only different patients but also among different lesions of the same patient. Invasive lesions of MPLC exhibited significantly higher TCR diversity and lower TCR expansion than those of SN. Intriguingly, different lesions of the same patient always shared a certain proportion of TCR clonotypes. Significant clonal expansion could be observed in shared TCR clonotypes, particularly in those existing in all lesions of the same patient. In conclusion, this study provided evidences of the distinctive mutational landscape, activation of oncogenic signaling pathways, and TCR repertoire in MPLC as compared with SN. The significant clonal expansion of shared TCR clonotypes demonstrated the existence of immune commonality among different lesions of the same patient and shed new light on the individually tailored precision therapy for MPLC.


Assuntos
Neoplasias Pulmonares , Mutação , Neoplasias Primárias Múltiplas , Receptores de Antígenos de Linfócitos T , Humanos , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Neoplasias Primárias Múltiplas/imunologia , Neoplasias Primárias Múltiplas/genética , Neoplasias Primárias Múltiplas/patologia , Masculino , Feminino , Pessoa de Meia-Idade , Idoso
4.
FASEB J ; 37(10): e23176, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37665592

RESUMO

Mycoplasma spp., the smallest self-replicating and genome-reduced organisms, have raised a great concern in both the medical and veterinary fields due to their pathogenicity. The molecular determinants of these wall-less bacterium efficiently use their limited genes to ensure successful infection of the host remain unclear. In the present study, we used the ruminant pathogen Mycoplasma bovis as a model to identify the key factors for colonization and invasion into host cells. We constructed a nonredundant fluorescent transposon mutant library of M. bovis using a modified transposon plasmid, and identified 34 novel adhesion-related genes based on a high-throughput screening approach. Among them, the ΔLppB mutant exhibited the most apparent decrease in adhesion to embryonic bovine lung (EBL) cells. The surface-localized lipoprotein LppB, which is highly conserved in Mycoplasma species, was then confirmed as a key factor for M. bovis adhesion with great immunogenicity. LppB interacted with various components (fibronectin, vitronectin, collagen IV, and laminin) of host extracellular matrix (ECM) and promoted plasminogen activation through tPA to degrade ECM. The 439-502 amino acid region of LppB is a critical domain, and F465 and Y493 are important residues for the plasminogen activation activity. We further revealed LppB as a key factor facilitating internalization through clathrin- and lipid raft-mediated endocytosis, which helps the Mycoplasma invade the host cells. Our study indicates that LppB plays a key role in Mycoplasma infection and is a potential new therapeutic and vaccine target for Mycoplasma species.


Assuntos
Mycoplasma bovis , Animais , Bovinos , Mycoplasma bovis/genética , Clatrina , Colágeno Tipo IV , Mutagênese , Plasminogênio
5.
Nanotechnology ; 35(4)2023 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-37852225

RESUMO

The morphology-dependent localized surface plasmon resonance of gold nanostructures has been widely utilized for designing sensors. One method relies on the color change of gold nanoparticles upon etching. In previous work, TMB2+oxidized from 3,3',5,5'-tetramethylbenzidine (TMB) was found to etch gold nanorods (AuNRs), leading to a spectrum of different colors. However, the preparation of TMB2+needs the addition of a strong acid and other harsh conditions. Herein, a new colorimetric biosensing platform was developed using urchin-like gold nanoparticles (AuNUs). Compared with AuNRs, the etching of AuNUs can happen under mild conditions by TMB+at pH 6, protecting enzymes and proteins from denaturation. The role of CTAB surfactant was dissected, and its bromide ions were found to be involved in the etching process. Based on these observations, a one-step colorimetric detection of H2O2was realized by using horseradish peroxidase and H2O2to oxidize TMB. Within 30 min, this system achieved a detection limit of 80 nM H2O2. This work offered fundamental insights into the etching of anisotropic gold nanostructures and optimized the etching conditions. These advancements hold promise for broader applications in biosensing and analytical chemistry.


Assuntos
Técnicas Biossensoriais , Nanopartículas Metálicas , Ouro/química , Colorimetria/métodos , Peróxido de Hidrogênio/química , Nanopartículas Metálicas/química , Técnicas Biossensoriais/métodos
6.
Bioorg Chem ; 136: 106543, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37119784

RESUMO

Curcumin is identified that it has the potential to treat Parkinson's disease (PD), but its instability limits its further application in clinic. The mono-carbonyl analogs of curcumin (MACs) with diketene structure can effectively improve its stability, but it is highly toxic. In the present study, a less cytotoxic and more stable monoketene MACs skeleton S2 was obtained, and a series of monoketene MACs were synthesized by combining 4-hydroxy-3­methoxy groups of curcumin. In the 6-OHDA-induced PD's model in-vitro, some compounds exhibited significant neurotherapeutic effect. The quantitative structure-activity relationship (QSAR) model established by the random forest algorithm (RF) for the cell viability rate of above compounds showed that the statistical results are good (R2 = 0.883507), with strong reliability. Among all compounds, the most active compound A4 played an important role in neuroprotection in the PD models both in vitro and in vivo by activating AKT pathway, and then inhibiting the apoptosis of cells caused by endoplasmic reticulum (ER) stress. In the PD model in-vivo, compound A4 significantly improved survival of dopaminergic neurons and the contents of neurotransmitters. It also enhanced the retention of nigrostriatal function which was better than the effect in the mice treated by Madopar, a classical clinical drug for PD. In summary, we screened out the compound A4 with high stability, less cytotoxic monoketene compounds. And these founding provide evidence that the compound A4 can protect dopaminergic neurons via activating AKT and subsequently suppressing ER stress in PD.


Assuntos
Curcumina , Fármacos Neuroprotetores , Doença de Parkinson , Animais , Camundongos , Apoptose , Curcumina/farmacologia , Curcumina/uso terapêutico , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Reprodutibilidade dos Testes
7.
Lipids Health Dis ; 22(1): 126, 2023 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-37563575

RESUMO

INTRODUCTION: Hearing loss is a prevalent health concern, and dietary factors, such as fatty acid intake, may play a role in its development. The current study aimed to investigate the association between the intake of dietary fatty acids and hearing thresholds among U.S. adults. METHODS: The researchers examined data from the National Health and Nutrition Examination Survey (NHANES), including 7,623 participants with available dietary fatty acid intake and audiometry data. Dietary fatty acid intake was assessed using dietary recalls, and hearing thresholds were measured using pure-tone audiometry. Multivariate linear regression models and smoothing curve fitting were utilized to explore the associations between dietary fatty acid intake and hearing thresholds, adjusting for relevant covariates. RESULTS: This study reveals a direct association between both low and high frequency pure tone average (PTA) hearing thresholds and the dietary intake of total saturated fatty acids (SFAs) and total polyunsaturated fatty acids (PUFAs). Conversely, the intake of total monounsaturated fatty acids (MUFAs) demonstrates an inverted U-shaped correlation with low-frequency and high-frequency PTA hearing thresholds, having inflection points at 11.91 (energy (%)) and 10.88 (energy (%)), respectively. CONCLUSION: Dietary intake of certain fatty acids may influence hearing thresholds in adults.


Assuntos
Gorduras na Dieta , Ácidos Graxos , Adulto , Humanos , Inquéritos Nutricionais , Ácidos Graxos Insaturados , Ácidos Graxos Monoinsaturados , Audição
8.
Molecules ; 28(18)2023 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-37764443

RESUMO

In fungi, the methylcitrate cycle converts cytotoxic propionyl-coenzyme A (CoA) to pyruvate, which enters gluconeogenesis. The glyoxylate cycle converts acetyl-CoA to succinate, which enters gluconeogenesis. The tricarboxylic acid cycle is a central carbon metabolic pathway that connects the methylcitrate cycle, the glyoxylate cycle, and other metabolisms for lipids, carbohydrates, and amino acids. Fungal citrate synthase and 2-methylcitrate synthase as well as isocitrate lyase and 2-methylisocitrate lyase, each evolved from a common ancestral protein. Impairment of the methylcitrate cycle leads to the accumulation of toxic intermediates such as propionyl-CoA, 2-methylcitrate, and 2-methylisocitrate in fungal cells, which in turn inhibits the activity of many enzymes such as dehydrogenases and remodels cellular carbon metabolic processes. The methylcitrate cycle and the glyoxylate cycle synergistically regulate carbon source utilization as well as fungal growth, development, and pathogenic process in pathogenic fungi.


Assuntos
Ciclo do Ácido Cítrico , Fungos , Acetilcoenzima A , Fungos/metabolismo , Carbono/metabolismo , Glioxilatos/metabolismo
9.
Zhongguo Yi Liao Qi Xie Za Zhi ; 47(5): 557-561, 2023 Sep 30.
Artigo em Chinês | MEDLINE | ID: mdl-37753897

RESUMO

With the highlighted advantages of 3D printing technology in the field of dental prosthodontics, there is increasing in the numbers of registration applications for additive manufacturing customized dentures. However, there is still a lack of unified analysis in the core elements of process control, the key points of registration and the safety production quality control. Based on the current research status of the industry, the study is intended to clarify confusion and difficulties, deeply analyse the mechanism of the product defects, sort the core elements of process control, then try to establish a systematic evaluation system from product performance research, key process verification, production quality control and the description of registration files, so that it can provide help for practitioners to clarify research direction, establishing quality management system, improving the efficiency of registration and ensuring product quality.

10.
Sensors (Basel) ; 22(7)2022 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-35408396

RESUMO

Effectively capturing and sensitively detecting cancer cells are critical to clinical diagnosis and cancer therapy. In this work, we prepared gold nanostar-decorated graphene oxide (GO-AuNSs) nanocomposites using a ultraviolet (UV)-induced strategy, and then modified them with a layer of bio-complex rBSA-FA (coupled reduced bovine serum albumin with folic acid) to generate GO-AuNSs@rBSA-FA nanocomposites. Herein, the application of GO and AuNSs not only strengthened the conductivity of the sensing platform but also guaranteed nanocomposites with biocompatible performance. Moreover, the adopted rBSA-FA layer could effectively enhance the stability and specificity towards gastric cancer cells (MGC-803). According to a systemic construction procedure, a novel electrochemical cytosensor based on GO-AuNSs@rBSA-FA was fabricated for MGC-803 cell detection. With the assistance of cyclic voltammetry (CV) and differential pulse voltammetry (DPV), the cytosensor reached a detection limit of 100 cell/mL in a wide linear range of 3 × 102~7 × 106 cell/mL towards MGC-803 cells. The good electrochemical characteristics for the cancer cell analysis indicate a promising prospect of this electrochemical cytosensor in clinical cancer diagnosis.


Assuntos
Técnicas Biossensoriais , Grafite , Nanocompostos , Neoplasias Gástricas , Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/métodos , Ouro , Humanos , Limite de Detecção , Neoplasias Gástricas/diagnóstico
11.
Int J Mol Sci ; 23(14)2022 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-35887015

RESUMO

Magnaporthe oryzae is the causal agent of rice blast, leading to significant reductions in rice and wheat productivity. Nap1 is a conserved protein in eukaryotes involved in diverse physiological processes, such as nucleosome assembly, histone shuttling between the nucleus and cytoplasm, transcriptional regulation, and the cell cycle. Here, we identified Nap1 and characterized its roles in fungal development and virulence in M. oryzae. MoNap1 is involved in aerial hyphal and conidiophore differentiation, sporulation, appressorium formation, plant penetration, and virulence. ΔMonap1 generated a small, elongated, and malformed appressorium with an abnormally organized septin ring on hydrophobic surfaces. ΔMonap1 was more sensitive to cell wall integrity stresses but more resistant to microtubule stresses. MoNap1 interacted with histones H2A and H2B and the B-type cyclin (Cyc1). Moreover, a nuclear export signal (NES) domain is necessary for Nap1's roles in the regulation of the growth and pathogenicity of M. oryzae. In summary, NAP1 is essential for the growth, appressorium formation, and pathogenicity of M. oryzae.


Assuntos
Magnaporthe , Oryza , Ascomicetos , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica , Histonas/metabolismo , Naftalenos , Proteína 1 de Modelagem do Nucleossomo/genética , Proteína 1 de Modelagem do Nucleossomo/metabolismo , Oligopeptídeos , Oryza/metabolismo , Doenças das Plantas/microbiologia , Esporos Fúngicos/metabolismo , Virulência
12.
Environ Monit Assess ; 194(11): 839, 2022 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-36169726

RESUMO

Suspended sediment concentration (c) has been considered a critical environmental factor in reef habitats; however, the values and variations of c are not evident in a unique reef mainly created by crustose coralline algal concretions compared to abundant studies in coral reefs. The results of real-time and long-term monitoring of waves and c over the intertidal algal reef are reported because of the construction of an offshore industrial harbor near the reef. The real-time monitoring systems were based on techniques, including optical backscatter sensors (OBSs) for measuring c, pressure sensors for measuring waves, data loggers, and wireless networks for data transmission. The instruments sampled every hour and ran continuously and automatically for years. The OBS measurement was compared and validated with biweekly water sampling. A good correlation between the results of the two methods was observed. Nevertheless, more calibrations of OBSs in different seasons reduced the variance between the two methods over a year-long timescale. The year-long data showed a remarkable seasonal variation in c. The average c was approximately 140 mg/l during the winter season, while it was only approximately 70 mg/l during the summer season. The observed c was higher than that in other coral reef environments; the elevated and highly variable c, ranging from approximately 0 to 500 mg/l, may be one factor that creates the unique algae reef environment. The year-long measurement of waves and c showed that the variation in c was mainly due to the variation in waves in different seasons and was well correlated with the wave-induced bed shear stress. The real-time and long-term data measured by the system will aid in better understanding and providing useful environmental data for accessing future environmental changes and protecting reef habitats.


Assuntos
Antozoários , Monitoramento Ambiental , Animais , Recifes de Corais , Ecossistema , Sedimentos Geológicos , Água
13.
Anal Chem ; 93(21): 7707-7713, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33999595

RESUMO

In a typical biosensor, a biomolecule such as an aptamer is used for target recognition, and a nanomaterial is used for signal generation. Herein, we communicate a reverse system using a nanomaterial for target recognition and a DNA for signaling. We discovered that a classic metal-organic framework material, zeolitic imidazolate framework (ZIF)-67, has ultrahigh selectivity for recognizing adenosine triphosphate (ATP), allowing a fluorescently labeled DNA oligonucleotide to be used for signal generation. This sensor showed up to a 24-fold increase in fluorescence upon adding 1 mM ATP, while the fluorescence increase after adding adenosine or guanosine triphosphate was less than twofold. Its selectivity is much better than that of the ATP aptamer, which binds adenosine even better. Using isothermal titration calorimetry, the selective binding of ATP was independently verified. This sensor has a detection limit of 29 nM ATP and it can even detect ATP in serum. By replacing Co2+ with Zn2+ to form ZIF-8 or by using CoO, the selectivity for ATP was lost. Therefore, by sophisticated material design, ultrahigh selectivity for molecular recognition can be achieved.


Assuntos
Aptâmeros de Nucleotídeos , Técnicas Biossensoriais , Estruturas Metalorgânicas , Zeolitas , Trifosfato de Adenosina
14.
Chemistry ; 27(37): 9597-9604, 2021 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-33857336

RESUMO

Oxidase-mimicking nanozymes are more desirable than peroxidase-mimicking ones since H2 O2 can be omitted. However, only a few nanomaterials are known for oxidase-like activities. In this work, we compared the activity of Mn2 O3 , Mn3 O4 and MnO2 and found that Mn2 O3 had the highest oxidase activity. Interestingly, the activity of Mn2 O3 was even inhibited by H2 O2 . The oxidase-like activity of Mn2 O3 was not much affected by the presence of proteins such as bovine serum albumin (BSA), but the physisorption of antibodies to Mn2 O3 was not strong enough to withstand the displacement by BSA. We then treated Mn2 O3 with 3-aminopropyltriethoxysilane to graft an amine group, which was used to conjugate antibodies using glutaraldehyde as a crosslinker. A one-step indirect competitive ELISA (icELISA) was developed for the detection of isocarbophos, and an IC50 of 261.7 ng/mL was obtained, comparable with the results of the standard two-step assay using horseradish peroxidase (HRP)-labeled antibodies. This assay has the advantage of significant timesaving for rapid detection of large amounts of samples. This work has discovered a highly efficient oxidase-mimicking nanozyme useful for various nano- and analytical applications.


Assuntos
Técnicas Biossensoriais , Oxirredutases , Compostos de Manganês , Óxidos , Peroxidase
15.
Langmuir ; 37(3): 1037-1044, 2021 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-33435677

RESUMO

Montmorillonite (MMT) is a two-dimensional (2D) clay material. Its abundance on the early earth has attracted studies for its role in prebiotic reactions, and adsorption of DNA to MMT is potentially important for understanding the origin of life. Although several possible models of DNA adsorption on MMT have been established, a consensus on the adsorption mechanism has yet to be formed, thereby a fundamental adsorption study is performed here. Adding up to 300 mM NaCl failed to promote DNA adsorption on MMT, Al2O3, or SiO2 nanoparticles. For polyvalent metals, DNA adsorption was achieved following the order Ce3+ > Cu2+ > Ni2+ > Zn2+. Among them, Ce3+ and Cu2+ inverted the surface charge of MMT to positive. In addition, using washing experiments, Cu2+- and Ce3+-mediated adsorption mainly depended on the DNA phosphate backbone, while Ni2+ and Zn2+ interacted with the backbone phosphate groups and adenine bases of DNA. Overall, these polyvalent metal ions promoted DNA adsorption via a cation bridge model. This research provides new insights into the surface interactions of MMT and DNA, which is conducive to future work on the interaction between clays and biopolymers.


Assuntos
Bentonita , Oligonucleotídeos , Adsorção , DNA , Íons , Dióxido de Silício
16.
J Org Chem ; 86(23): 17371-17379, 2021 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-34783555

RESUMO

The spiro scaffold chiral organocatalyst of 3,2'-pyrrolidinyl spiro-oxindole amine was successfully prepared from racemic spiro-oxindole amine using l-menthol as a chiral pool in 4 steps in 28%-40% overall yields with at least 99% ee in scale-up preparation, and its catalytic activity was evaluated in the enantioselective aldol condensation between 3-(3-hydroxy-1H-pyrazol-1-yl)-oxindole and paraformaldehyde. The spiro organocatalyst showed superior catalytic activity and selectivity compared with its counterparts, and most substrates offered good to excellent results with up to 96% yield in 96% ee.


Assuntos
Compostos de Espiro , Aldeídos , Aminas , Formaldeído , Indóis , Estrutura Molecular , Oxindóis , Polímeros , Estereoisomerismo
17.
Analyst ; 146(3): 864-873, 2021 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-33231579

RESUMO

Having the benefits of low cost, excellent stability and tolerance to extreme conditions, nanozymes are a potential alternative of horseradish peroxidase (HRP) or other enzymes for bioanalytical chemistry, especially immunoassays. CeO2 nanoparticles have oxidase-mimicking activity and can avoid the use of unstable H2O2. For robust assays, the effect of proteins on the activity of CeO2 needs to be carefully studied. Herein, we studied the adsorption and desorption of bovine serum albumin (BSA) from CeO2. The CeO2 nanoparticles exhibited a higher protein adsorption capacity compared to the other tested metal oxide nanoparticles. Although the oxidase-like activity of CeO2 was inhibited by BSA, low concentrations of phosphate and fluoride ions boosted the activity of protein-capped CeO2. CeO2 was still active under strong acidic conditions and at high temperature, while HRP lost its activity. For immunoassay development, we covered CeO2 with an amine-modified silane for covalent conjugation to antibodies. A one-step indirect competitive ELISA for fenitrothion was developed, and an IC50 value of 35.6 ng mL-1 and a limit of detection of 2.1 ng mL-1 were obtained.


Assuntos
Cério , Nanopartículas Metálicas , Peróxido de Hidrogênio , Imunoensaio , Oxirredutases
18.
Angew Chem Int Ed Engl ; 60(23): 12985-12991, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33792133

RESUMO

Acrydite-modified DNA is the most frequently used reagent to prepare DNA-functionalized hydrogels. Herein, we show that unmodified penta-adenine (A5 ) can reach up to 75 % conjugation efficiency in 8 h under a freezing polymerization condition in polyacrylamide hydrogels. DNA incorporation efficiency was reduced by forming duplex or other folded structures and by removing the freezing condition. By designing diblock DNA containing an A5 block, various functional DNA sequences were attached. Such hydrogels were designed for ultrasensitive DNA hybridization and Hg2+ detection, with detection limits of 50 pM and 10 nM, respectively, demonstrating the feasibility of using unmodified DNA to replace acrydite-DNA. The same method worked for both gel nanoparticles and monoliths. This work revealed interesting reaction products by exploiting freezing and has provided a cost-effective way to attach DNA to hydrogels.


Assuntos
Adenina/química , DNA/análise , Congelamento , Hidrogéis/química , Mercúrio/análise , Nanopartículas/química , Estrutura Molecular
19.
Anal Chem ; 92(13): 9370-9378, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32515584

RESUMO

Target-directed aptamer adsorption by gold nanoparticles (AuNPs) has been widely used to develop label-free colorimetric biosensors. However, the potential interactions between target molecules and AuNPs have not been considered, which may lead to misinterpretation of analytical results. In this work, the detection of dopamine, melamine, and K+ was studied as model systems to address this problem. First, dopamine and two control molecules all induced the aggregation of citrate-capped AuNPs with apparent Kd's of 5.8 µM dopamine, 51.6 µM norepinephrine, and 142 µM tyramine. Isothermal titration calorimetry measured the aptamer Kd to be 1.9 µM dopamine and 16.8 µM norepinephrine, whereas tyramine cannot bind. Surface enhanced Raman spectroscopy confirmed direct adsorption of dopamine, and the adsorbed dopamine inhibited the adsorption of DNA. Using a typical salt-induced colorimetric detection protocol, a similar color response was observed regardless of the sequence of DNA, indicating the observed color change reflected the adsorption of dopamine by the AuNPs instead of the binding of dopamine by the aptamer. For this label-free sensor to work, the interaction between the target molecule and AuNPs should be very weak, while dopamine represents an example of strong interactions. For the other two systems, the melamine detection did not reflect aptamer binding either but the K+ detection did, suggesting melamine also strongly interacted with AuNPs, whereas K+ had very weak interactions with AuNPs. Since each target molecule is different, such target/AuNP interactions need to be studied case-by-case to ensure the sensing mechanism.


Assuntos
Aptâmeros de Nucleotídeos/metabolismo , Técnicas Biossensoriais/métodos , Dopamina/análise , Nanopartículas Metálicas/química , Triazinas/análise , Adsorção , Aptâmeros de Nucleotídeos/química , Ácido Cítrico/química , Colorimetria , DNA/química , Ouro/química
20.
Langmuir ; 36(45): 13708-13715, 2020 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-33161721

RESUMO

Finding DNA sequences that can adsorb strongly on nanomaterials is critical for bioconjugate and biointerface chemistry. In most previous work, unmodified DNA with a phosphodiester backbone (PO DNA) were screened or selected for adsorption on inorganic surfaces. In this work, the adsorption of phosphorothioate (PS)-modified DNA (PS DNA) on graphene oxide (GO) is studied. By use of fluorescently labeled oligonucleotides as probes, all the tested PS DNA strands are adsorbed more strongly on GO compared to the PO DNA of the same sequence. The adsorption mechanism is probed by washing the adsorbed DNA with proteins, surfactants, and urea. Molecular dynamics simulations show that van der Waals forces are responsible for the tighter adsorption of PS DNA. Polycytosine (poly-C) DNA, in general, has a high affinity for the GO surface, and PS poly-C DNA can adsorb even stronger, making it an ideal anchoring sequence on GO. With this knowledge, noncovalent functionalization of GO with a diblock DNA is demonstrated, where a PS poly-C block is used to anchor on the surface. This conjugate achieves better hybridization than the PO DNA of the same sequence for hybridization with the complementary DNA.


Assuntos
Grafite , Oligonucleotídeos Fosforotioatos , Adsorção , DNA
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