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1.
Mikrochim Acta ; 191(1): 78, 2024 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-38182922

RESUMO

New portable hydrogel sensors for Al3+ and Fe3+ detection were designed based on the aggregation-induced emission (AIE) and color change of N-doped carbon dots (N-CDs). N-CDs with yellow fluorescence were prepared by a one-pot hydrothermal method from 2,5-dihydroxyterephthalic acid and acrylamide. The fluorescence of N-CDs was enhanced by Al3+ about 20 times and quenched by Fe3+. It was interesting that although Fe3+ showed obvious quenching on the fluorescence of N-CDs it did not cause a noticeable change in the fluorescence of N-CDs + Al3+. The colorless solution of N-CDs appeared blue in the presence of Fe3+ without the influence of Al3+. Therefore, the turn-on fluorometry and colorimetry systems based on N-CDs were constructed for the simultaneous detection of Al3+ and Fe3+. Furthermore, the portable sensing of Al3+ and Fe3+ was realized with the assistance of hydrogel, filter paper, cellulose acetate, and cellulose nitrate film. The proposed approach was successfully applied to the detection of Al3+ and Fe3+ in food samples and cell imaging.

2.
Anal Chem ; 95(27): 10353-10361, 2023 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-37339258

RESUMO

Pancreatic cancer patients predominantly present with advanced disease at diagnosis, contributing to its high mortality. A noninvasive, fast screening method to detect this disease is an unmet need. Tumor-derived extracellular vesicles (tdEVs) bearing information from parental cells have emerged as a promising cancer diagnostic biomarker. However, most tdEV-based assays have impractical sample volumes and time-consuming, complex, and costly techniques. To overcome these limitations, we developed a novel diagnostic method for pancreatic cancer screening. Our approach utilizes the mitochondrial DNA to nuclear DNA ratio of EVs as a collective cell-specific characteristic. We introduce EvIPqPCR, a fast method that combines immunoprecipitation (IP) and qPCR quantification to detect tumor-derived EVs directly from serum. Importantly, our method employs DNA isolation-free and duplexing probes for qPCR, saving at least 3 h. This technique has the potential to serve as a translational assay for cancer screening with a weak correlation to prognosis biomarkers and sufficient discriminatory power among healthy controls, pancreatitis, and pancreatic cancer cases.


Assuntos
Vesículas Extracelulares , Neoplasias Pancreáticas , Humanos , Linhagem Celular Tumoral , Neoplasias Pancreáticas/diagnóstico , Biomarcadores Tumorais , Neoplasias Pancreáticas
3.
Anal Chem ; 95(48): 17945-17953, 2023 12 05.
Artigo em Inglês | MEDLINE | ID: mdl-38000786

RESUMO

N6-Methyladenosine (m6A) is the most pervasive and evolutionarily conserved epitranscriptomic modification in long noncoding RNA (lncRNA), and its dysregulation may induce aberrant transcription and translation programs. Herein, we demonstrate the methylation-powered assembly of a single quantum dot (QD)-based fluorescence resonance energy transfer (FRET) nanosensor for antibody- and enzyme-free monitoring of locus-specific m6A in clinical tissues. The m6A-sensitive DNAzyme VMC10 is employed to identify a specific m6A site in lncRNA, and it catalyzes the hydrolytic cleavage of unmethylated lncRNA. The cleaved lncRNA fails to trigger the subsequent catalytic hairpin assembly (CHA) reaction due to the energy barrier. In contrast, when m6A-lncRNA is present, the methyl group in m6A protects lncRNA from VMC10-mediated cleavage. With the aid of an assistant probe, the retained intact m6A-lncRNA is released from the VMC10/lncRNA complex and subsequently triggers the CHA reaction, generating abundant AF647/biotin dual-labeled duplexes. The assembly of AF647/biotin dual-labeled duplexes onto 605QD results in efficient FRET between 605QD and AF647. The FRET signal can be simply quantified by single-molecule detection. Notably, this assay can be implemented in an antibody-free and enzyme-free manner. This nanosensor can sensitively quantify target m6A with a detection limit of 0.47 fM, and it can discriminate as low as a 0.001% m6A level from excess coexisting counterparts. Importantly, this nanosensor can monitor the cellular m6A level with single-cell sensitivity and profile target m6A expression in breast cancer and healthy para-cancerous tissues, providing a powerful tool for studying the physiological and pathological functions of m6A.


Assuntos
Técnicas Biossensoriais , Pontos Quânticos , RNA Longo não Codificante , Transferência Ressonante de Energia de Fluorescência/métodos , Metilação , Biotina , RNA Longo não Codificante/genética , Anticorpos
4.
Small ; 19(2): e2204298, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36354195

RESUMO

Sensitive detection of extracellular vesicles (EVs) as emerging biomarkers has shown great promises for disease diagnosis. Plasmonic metal nanostructures conjugated with molecules that bind specific biomarker targets are widely used for EVs sensing but involve tradeoffs between particle-size-dependent signal intensity and conjugation efficiency. One solution to this problem would be to induce nucleation on nanoparticles that have successfully bound a target biomarker to permit in situ nanoparticle growth for signal amplification, but approaches that are evaluated to date require harsh conditions or lack nucleation specificity, prohibiting their effective use with most biological specimens. This study describes a one-step in situ strategy to induce monocrystalline copper shell growth on gold nanorod probes without decreasing signal by disrupting probe-target interactions or lipid bilayer integrity to enable EV biomarker detections. This approach increases the detected nanoparticle signal about two orders of magnitude after a 10 min copper nanoshell growth reaction. This has significant implications for improved disease detection, as indicated by the ability of a novel immunoassay using this approach to detect low abundance EVs carrying a pathogen-derived biomarker, after their direct capture from serum, to facilitate the diagnosis of tuberculosis cases in a diagnostically challenging pediatric cohort.


Assuntos
Vesículas Extracelulares , Nanopartículas , Humanos , Criança , Cobre/metabolismo , Biomarcadores/análise , Bicamadas Lipídicas/metabolismo , Vesículas Extracelulares/metabolismo
5.
Analyst ; 148(8): 1653-1671, 2023 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-36960759

RESUMO

With the global penetration of skin care awareness and upgrading of personal care awareness, the use rate of cosmetics and personal skin care products has been increasing worldwide. It is particularly important to monitor the quality and safety of skin cosmetics. In accordance with the requirements of the 7th Amendment of the European Cosmetics Directive 1223/2009, in vitro test methods have been developed to replace animal experiments, such as the 2D test, 3D test, microfluidic skin chip test, etc. The microfluidic skin chip overcomes the shortcomings of the 2D test and the 3D test that lack the complexity of human skin through fine control of the human skin microenvironment and induction of relevant mechanical stimulation. High similarity to real human skin through simulation of the vascular system and immune response. Therefore, the microfluidic skin chip is considered as a valuable and effective tool for the in vitro screening of cosmetics. In this paper, we reviewed the detection methods and technologies of common chemical substances, toxic elements, active substances and adverse reactions in vitro in quality monitoring of cosmetics. The most advantageous microfluidic skin chip technology is also introduced. The material and technology progress of skin chips used in cosmetic screening is reviewed and discussed. Then the application of microfluidic design in cosmetic screening in vitro is summarized.


Assuntos
Cosméticos , Microfluídica , Animais , Humanos , Microfluídica/métodos , Pele
6.
Mikrochim Acta ; 190(7): 260, 2023 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-37318602

RESUMO

High-throughput screening platforms are fundamental for the rapid and efficient processing of large amounts of experimental data. Parallelization and miniaturization of experiments are important for improving their cost-effectiveness. The development of miniaturized high-throughput screening platforms is essential in the fields of biotechnology, medicine, and pharmacology. Currently, most laboratories use 96- or 384-well microtiter plates for screening; however, they have disadvantages, such as high reagent and cell consumption, low throughput, and inability to avoid cross-contamination, which need to be further optimized. Droplet microarrays, as novel screening platforms, can effectively avoid these shortcomings. Here, the preparation method of the droplet microarray, method of adding compounds in parallel, and means to read the results are briefly described. Next, the latest research on droplet microarray platforms in biomedicine is presented, including their application in high-throughput culture, cell screening, high-throughput nucleic acid screening, drug development, and individualized medicine. Finally, the challenges and future trends in droplet microarray technology are summarized.


Assuntos
Ensaios de Triagem em Larga Escala , Ensaios de Triagem em Larga Escala/métodos , Avaliação Pré-Clínica de Medicamentos , Análise em Microsséries/métodos
7.
Analyst ; 147(15): 3444-3450, 2022 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-35792747

RESUMO

Heat shock protein 70 (Hsp70), belonging to the heat shock protein (HSP) family, is reported to be a potential diagnostic biomarker. In this work, a lateral flow immunostrip was fabricated for the sensitive and rapid determination of Hsp70 by the incorporation of fluorescence and upconversion nanoparticle probes. The upconversion nanoparticles (UCNPs, size ∼39 nm, λex = 980 nm; λem = 540 nm) consisting of a NaYF4:Yb/Er core and polyacrylic acid-modified shell were covalently coupled with Hsp70 antibodies to form the signal probe, which was characterized by dynamic light scattering and zeta potential analyses. The lateral flow assay (LFA) was constructed based on the sandwich-type immunoassay using a sample pad, a test pad, and an adsorption pad on a PVP backing. Hsp70 antibody, IgG antibody and the signal probe were separately dropped on the test zone, the control zone of the test pad, and the sample pad, respectively. In the sandwich LFA, since two antibodies bind to Hsp70 antigenic epitopes, i.e. specific binding, it provided superior specificity and high sensitivity, making it an ideal sensing platform for complex samples like serum Hsp70 samples. The important parameters for the preparation of the lateral flow immunostrips were optimized. Under the optimized conditions, Hsp70 can be detected using the increased fluorescence intensity of UCNPs with a wide linear range from 0.11 to 12 ng mL-1, low detection limit of 0.06 ng mL-1, small sample volume (120 µL), short assay time (15 min) and good reproducibility. The fluorescence method was successfully applied in the determination of Hsp70 in serum samples with good recovery. By combining the accessibility of the lateral flow immunostrips and upconversion nanoparticles, the fluorescence method can serve as a point-of-care testing method for protein assays with high sensitivity and fast detection.


Assuntos
Corantes Fluorescentes , Nanopartículas , Anticorpos , Proteínas de Choque Térmico HSP70 , Imunoensaio/métodos , Nanopartículas/química , Reprodutibilidade dos Testes
8.
Sens Actuators A Phys ; 3462022 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-37273787

RESUMO

Extracellular vesicles (EVs) bearing biomolecules from parental cells can represent a novel source of disease biomarkers and are under intensive study for their clinical potential. Tunable resistive pulse sensing (TRPS) quantifies the magnitude of a small ionic resistive pulse current to determine the size, concentration, and zeta potential of EVs. Environmental noise is a common limiting factor that affects the precision of sensing devices. TRPS is particularly vulnerable to environmental noise, including both mechanical and electrical. The upper detection limit of the TRPS relies on the physical size of the elastomeric tunable nanopore. The lower limit relies on the electrical signal-to-noise ratio. Guided by simulation, we designed an external device to suppress environmental noise for TRPS measurement. Both mechanical and electrical environmental noise reductions were observed after using the shield. The study also validated the noise reduction function of the shield by quantifying EVs from different cell origins. Detection of EVs smaller than 200 nm was improved by using the shield; which was reported challenging for conventional quantification methods. The study highlighted a feasible approach to solve environmental noise challenges for TRPS based EV quantification.

9.
Sens Actuators B Chem ; 3382021 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-33927481

RESUMO

Alzheimer's disease and other neurodegenerative disorders are becoming more prevalent as advances in technology and medicine increase living standards and life expectancy. Alzheimer's disease is thought to initiate development early in the patient's life and progresses continuously into old age. This process is characterized molecularly by the amyloid hypothesis, which asserts that self-aggregating amyloid peptides are core to the pathophysiology in Alzheimer's progression. Precise quantification of amyloid peptides in human bodily fluid samples (i.e. cerebrospinal fluid, blood) may inform diagnosis and prognosis, and has been studied using established biosensing technologies like liquid chromatography, mass spectrometry, and immunoassays. However, existing methods are challenged to provide single molecule, quantitative analysis of the disease-causing aggregation process. Ultra-sensitive nanopore biosensors can step in to fill this role as a dynamic mapping tool. The work in this paper establishes characteristic signals of ß-amyloid 40 monomers, oligomers, and soluble aggregates, as well as a proof-of-concept foundation where a biological nanopore biosensor is used to monitor the extent of in vitro ß-amyloid 40 peptide aggregation at the single molecule level. This foundation allows for future work to expand in drug screening, diagnostics, and aggregation dynamic experiments.

10.
Nanomedicine ; 32: 102339, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33227538

RESUMO

MicroRNA (miRNA) has emerged as a promising genetic marker for cancer diagnosis and therapy because its expression level is closely related to the progression of malignant diseases. Herein, a label-free and selective fluorescence platform was proposed for miRNA based on light-up "G-quadruplex nanostring" via duplex-specific nuclease (DSN) mediated tandem rolling circle amplification (RCA). First, a long DNA generated from upstream RCA was designed with the antisense sequences for miR-21 and downstream RCA primer. Upon recognizing miR-21, the resulting DNA-RNA permitted DSN digestion and triggered downstream two-way RCA, and generation of abundant "G-quadruplex nanostring" binding with ZnPPIX for label-free fluorescent responses. In our strategy, the strong preference of DSN for perfectly matched DNA/RNA ensures its excellent selectivity. The developed method generated wide linear response with LOD of 1.019 fM. Additionally, the miR-21 levels in cell extracts have been evaluated, revealing the utility of this tool for biomedical research and clinical diagnosis.


Assuntos
Endonucleases/metabolismo , Quadruplex G , MicroRNAs/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Estudos de Viabilidade , Células HeLa , Humanos , MicroRNAs/metabolismo , Espectrometria de Fluorescência
11.
Mikrochim Acta ; 188(10): 352, 2021 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-34554325

RESUMO

Extracellular ATP as a purinergic signaling molecule, together with ATP receptor, are playing an important role in tumor growth, therapy resistance, and host immunity suppression. Meanwhile ATP is a crucial indicator for cellular energy status and viability, thus a vital variable for tissue regeneration and in vitro tissue engineering. Most recent studies on COVID-19 virus suggest infection caused ATP deficit and release as a major characterization at the early stage of the disease and major causes for disease complications. Thus, imaging ATP molecule in both cellular and extracellular contexts has many applications in biology, engineering, and clinics. A sensitive and selective fluorescence "signal-on" probe for ATP detection was constructed, based on the base recognition between a black hole quencher (BHQ)-labeled aptamer oligonucleotide and a fluorophore (Cy5)-labeled reporter flare. The probe was able to detect ATP in solution with single digit µM detection limit. With the assistance of lipofectamine, this probe efficiently entered and shined in the model cells U2OS within 3 h. Further application of the probe in specific scenery, cardio-tissue engineering, was also tested where the ATP aptamer complex was able to sense cellular ATP status in a semi-quantitative manner, representing a novel approach for selection of functional cardiomyocytes for tissue engineering. At last a slight change in probe configuration in which a flexible intermolecular A14 linker was introduced granted regeneration capability. These data support the application of this probe in multiple circumstances where ATP measurement or imaging is on demand.


Assuntos
Trifosfato de Adenosina/análise , Aptâmeros de Nucleotídeos , Carbocianinas , Corantes Fluorescentes , Animais , Animais Recém-Nascidos , Linhagem Celular , Fluorescência , Humanos , Miócitos Cardíacos , Ratos
12.
Anal Chem ; 92(2): 2244-2250, 2020 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-31874559

RESUMO

In the fabrication of cardiac tissue, an important factor is continuous measurement of its contraction features. A module that allows for a dynamic system capable of noninvasive and label-free monitoring of the contraction profile under administering chemicals and drugs is highly valuable for understanding accurate tissue mechanobiology. In this research, we have successfully demonstrated the use of surface plasmon resonance (SPR) technology for the first time to characterize the contractility of cardiac cells in response to Blebbistatin and ATP drug exposure in real tme. An optimal flow rate of 10 µL/min was selected for a continuous flow of warm media,and 10 µM drug administration effect was detected with high spatiotemporal sensitivity on contracting cardiomyocytes. Our drug screening has identified the source of the SPR periodic signal to be direct cell contraction rather than action potentials or calcium signaling. Per our results, SPR has high potential in applications in least-interference real-time and label-free tissue characterizations and cellular properties analysis from a functional and structural point of view.


Assuntos
Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Animais , Células Cultivadas , Compostos Heterocíclicos de 4 ou mais Anéis/administração & dosagem , Ratos , Ressonância de Plasmônio de Superfície , Fatores de Tempo
13.
Analyst ; 145(22): 7448, 2020 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-32966362

RESUMO

Correction for 'A facile deoxyuridine/biotin-modified molecular beacon for simultaneous detection of proteins and nucleic acids via a label-free and background-eliminated fluorescence assay' by Fei Yin, et al., Analyst, 2019, 144, 5504-5510, DOI: 10.1039/C9AN01016E.

14.
Analyst ; 144(18): 5504-5510, 2019 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-31389925

RESUMO

Simultaneous detection of different types of cancer biomarkers (nucleic acids and proteins) could facilitate early diagnosis of cancer and clinical treatment. Herein, a simultaneous detection platform of proteins and nucleic acids has been developed using a single substrate probe combining a label-free and background-eliminated fluorescence assay. Telomerase and telomerase RNA (TR) were chosen as the models. The molecular beacon (dU-BIO-HP) that contains deoxyuridine/biotin in its side arm, a TR recognition sequence in the loop and a telomerase substrate primer at the stem end was ingeniously designed. In the presence of telomerase, the stem of dU-BIO-HP is elongated by the addition of telomere repeats complementary to the assistant DNA. Furthermore, the formed dsDNA performed as engaging primers to initiate a SDA reaction, generating abundant G-quadruplex monomers. Similarly, on TR, the hybridization between TR and dU-BIO-HP can open its stem, triggering another SDA reaction, producing abundant short ssDNAs. With the G-quadruplex binding with ZnPPIX and ssDNA binding with SG for specific fluorescence responses, the label-free multiple detection can be achieved. In our strategy, the deoxyuridine of dU-BIO-HP acts as a barrier to block the DNA extension due to its strong inhibitory effects on DNA polymerase activity and to make sure that the two SDA reactions occurred independently. The biotin of dU-BIO-HP enables the reduction of the background from the binding between SG, ZnPPIX and dU-BIO-HP through streptavidin-biotin interaction. This method showed an excellent sensitivity with telomerase and TR detection limit of 2.18 HeLa cells per mL and 0.16 × 10-12 M, respectively. Furthermore, the telomerase and TR in different cell lines have been evaluated as powerful tools for biomedical research and clinical diagnosis.


Assuntos
Biomarcadores Tumorais/análise , RNA/análise , Telomerase/análise , Biotina/química , Linhagem Celular Tumoral , Sondas de DNA/química , Sondas de DNA/genética , Desoxiuridina/química , Ensaios Enzimáticos/métodos , Corantes Fluorescentes/química , Humanos , Limite de Detecção , Técnicas de Amplificação de Ácido Nucleico , Hibridização de Ácido Nucleico , Compostos Orgânicos/química , RNA/genética , Espectrometria de Fluorescência , Telomerase/genética
15.
Analyst ; 144(9): 2994-3004, 2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-30892312

RESUMO

The human telomerase reverse transcriptase catalytic subunit (hTERT) is the rate-limiting subunit of the telomerase holoenzyme. Down-regulating the expression of hTERT mRNA by antisense oligonucleotides would reduce the expression of hTERT, inhibit telomerase activity, and impair the growth of cancer cells in vitro. In this work, we propose a locked nucleic acid-functionalized gold nanoparticle flare probe (AuNP-probe). After transferring these probes into cells by endocytosis of the gold nanoparticles, the binding process of the antisense locked nucleic acid with hTERT mRNA along with gene regulation can be visualized by fluorescence recovery of flare-sequences. A significant decline in hTERT mRNA levels and the hTERT content occurred in cancer cells after treatment with the AuNP-probes, and only approximately 25% of the original level of hTERT mRNA remained after 72 h. AuNP-probe treated cancer cells were arrested in the G1 phase of the cell cycle and underwent apoptosis; cell viability decreased obviously compared with that of telomerase-negative normal cells.


Assuntos
DNA/química , Corantes Fluorescentes/química , Ouro/química , Nanopartículas Metálicas/química , RNA Mensageiro/metabolismo , Telomerase/metabolismo , Antineoplásicos/química , Antineoplásicos/farmacologia , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Carbocianinas/química , Catequina/análogos & derivados , Catequina/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , DNA/toxicidade , Regulação para Baixo , Inibidores Enzimáticos/farmacologia , Fluorescência , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Nanopartículas Metálicas/toxicidade , Microscopia Confocal/métodos , Microscopia de Fluorescência/métodos , Hibridização de Ácido Nucleico , Oligonucleotídeos/química , Oligonucleotídeos/genética , Oligonucleotídeos/toxicidade , Oligonucleotídeos Antissenso/química , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/toxicidade , RNA Mensageiro/genética , Telomerase/antagonistas & inibidores , Telomerase/genética , Fatores de Tempo
16.
Mikrochim Acta ; 186(9): 640, 2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31440852

RESUMO

A fluorometric method is described for "turn-on" sensing of pH values via black phosphorus quantum dots (BPQD). Water-stable BPQD were synthesized by a liquid exfoliation method and characterized by TEM, FT-IR, XPS, and absorption and fluorescence spectra. The nanoparticles of BPQD have a uniform distribution with an average size of 5.2 nm. They exhibit bright green fluorescence, with excitation/emission maxima at 420/515 nm. The fluorescence of the BPQD is likely to arise from the quasi-molecular fluorophores of polycyclic aromatic compounds carrying P-P, P-O-P, and PxOy functions on its surface. The protonation and deprotonation of hydroxyl groups of BPQD causes a different degree of quenching of the BPQD. At pH values below 4.0, protons bind to BPQD to form non-fluorescent ground state complexes. At pH values above 4.0, the hydroxyl groups become deprotonated, and this induces the recovery of fluorescence. The sensor has a linear response in the pH range of 1.0-9.0. It was successfully applied to the determination of the pH values in human urine and serum samples. Graphical abstract Schematic representation of the preparation of black phosphorus quantum dots (BPQDs) from powdered BP crystals using liquid-phase exfoliation in N-methyl-2-pyrrolidone solution. The BPQDs display green fluorescence at high pH values but no fluorescence at very low pH values.

17.
Analyst ; 143(7): 1593-1598, 2018 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-29517783

RESUMO

We developed a novel approach to determine formamidopyrimidine DNA glycosylase (FPG) activity by taking advantage of target-induced self-primed rolling circle amplification (RCA) and magnetic nanoprobes. Herein, a unique nick (8-oxoguanine, 8-oxoG) was positioned in duplex DNA containing P-circle and P1, which together serve as a FPG substrate, RCA template, and RCA primer probe. The presence of FPG specifically binds 8-oxoG and cleaves the P-circle into two parts, producing 5'-phosphoryl termini. A phosphodiester bond between the 5'-phosphoryl and 3'-hydroxyl termini was formed with the addition of T4 DNA ligase, producing an unnicked circular strand. Using the unnicked strand as the RCA template, the P1 hybridized with the circle probe as a primer will trigger the RCA process. The RCA reaction produces amounts of long tandem-repeat DNA tiles with multiple recognizing regions for the FAM modified DNA probes (FP) and biotin-modified DNA probes (BP). With the streptavidin-biotin interaction, the BPs and FPs can be easily immobilized on the surface of streptavidin-modified magnetic microbeads (MBs). Due to the RCA enhanced and highly-concentrated fluorescence accumulation on the MBs, an ultralow detection limit of 1.033 U mL-1 for FPG was obtained. Combined with the high tolerance capability of human blood serum owing to magnetic isolation, the FPG assays in human blood serum were also obtained using fluorescence and confocal laser scanning microscopy. These results indicate that this robust self-primed RCA combined with magnetic nanoprobes is an excellent candidate for quantitatively monitoring the FPG activity responsible for DNA oxidative damage-related clinical diagnosis and therapy.


Assuntos
DNA-Formamidopirimidina Glicosilase/análise , Nanopartículas , Técnicas de Amplificação de Ácido Nucleico , Sondas de DNA , DNA-Formamidopirimidina Glicosilase/sangue , Humanos , Magnetismo
18.
Lipids Health Dis ; 17(1): 251, 2018 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-30400958

RESUMO

BACKGROUND: Bitter melon (BM, Momordica charantia) has been accepted as an effective complementary treatment of metabolic disorders such as diabetes, hypertension, dyslipidemia and etc. However it is unclear whether BM can prevent the progression of atherosclerosis. To confirm the effects of BM on atherosclerosis and explore its underlying mechanisms, we design this study. METHODS: Twenty four male apolipoprotein E knock-out (ApoE-/-) mice aged 8 weeks were randomly divided into control group fed with high fat diet (HFD) only and BM group fed with HFD mixed with 1.2%w/w BM. After 16 weeks, body weight, food intake, blood glucose, serum lipids were measured and the atherosclerotic plaque area and its histological composition were analyzed. The expression of vascular cell adhesive molecules and inflammatory cytokines in the aortas were determined using quantitative polymerase chain reaction. RESULTS: Body weight gain and serum triglycerides (TG) significantly decreased in BM group. BM reduced not only the atherosclerotic plaque area and the contents of collagen fibers in atherosclerotic plaques but also the serum soluble vascular cell adhesion molecule (VCAM)-1 and P-selectin levels, as well as the expressions of monocyte chemoattractant protein (MCP)-1 and interleukin (IL)-6 in aortas. CONCLUSION: Our study indicates that dietary BM can attenuate the development of atherosclerosis in ApoeE-/- mice possibly through reducing triglyceride and anti-inflammation mechanism.


Assuntos
Aterosclerose/tratamento farmacológico , Hipertrigliceridemia/tratamento farmacológico , Inflamação/tratamento farmacológico , Momordica charantia/química , Preparações de Plantas/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Aterosclerose/prevenção & controle , Citocinas/sangue , Dieta Hiperlipídica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Triglicerídeos/sangue
19.
Mikrochim Acta ; 185(12): 550, 2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30443788

RESUMO

N/S/P-codoped carbon dots (CDs) are shown to be a viable fluorescent probe in a turn-off-on fluorometric assay for hydroquinone (HQ). The preparation of CDs was carried out using a one-step hydrothermal reaction starting with glyoxal and isocarbophos. The method is based on the formation of ground state complexes between CD and Fe(III) which leads to quenching of blue fluorescence (with excitation/emission peaks at 363/448 nm). On addition of HQ, it will be oxidized by Fe(III) upon which fluorescence recovers. This turn-off-on system can be utilized to quantify HQ. A linear relationship exists between fluorescence recovery and HQ concentration in range between 0.56 and 375 µM. The limit of detection is 0.16 µM. The assay was successfully applied to the determination of HQ in spiked water samples and developer samples. Graphical abstract Fluorometric determination of hydroquinone (with good selectivity over catechol and resorcinol) by using blue-emitting N/S/P-codoped carbon dots and the quenching effect of Fe(III).

20.
Mikrochim Acta ; 185(9): 419, 2018 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-30121832

RESUMO

An aptamer based method is described for the determination of 8-hydroxy-2'-deoxyguanosine (8-OHdG) using resonance light scattering (RLS). Magnetic nanoparticles (MNPs) were employed as RLS probes. The probe DNA was placed on the surface of MNPs, which produces a rather low RLS signal. If, however, probe DNA hybridizes with the aptamer against 8-OHdG, a sandwich structure will be formed. This results in a significant enhancement of RLS intensity. The aptamer was used as the recognition element to capture 8-OHdG. 8-OHdG has a stronger affinity for the aptamer than probe DNA, and the conformation of the aptamer therefore switches from a double-stranded to a G-quadruplex structure. As a result, MNPs labeled with probe DNA are released, and RLS intensity decreases. The method allows 8-OHdG to be detected with a linear response in the 32 pM - 12.0 nM concentration range and an 11 pM limit of detection (at 3.29SB/m, according to the recent recommendation of IUPAC). The MNPs can be reused 5 times by applying an external magnetic field for collection. The method was successfully applied to analyze human urine samples for its content of 8-OHdG. It was also found that the levels of 8-OHdG noticeably increased with the increase of the Air Quality Index. Conceivably, the method is a viable tool to investigate the relationship between 8-OHdG levels and the effect of air pollution. Graphical abstract A reusable sensing strategy was constructed to detect urinary 8-OHdG based on "turn-off" resonance light scattering. The LOD was as low as 11 pM. This study showed some preliminary data for the association between oxidative stress and air pollution.

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