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1.
Biosens Bioelectron ; 169: 112578, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32911317

RESUMO

The ongoing global pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to active research in its associated diagnostics and medical treatments. While quantitative reverse transcription polymerase chain reaction (qRT-PCR) is the most reliable method to detect viral genes of SARS-CoV-2, serological tests for specific antiviral antibodies are also important as they identify false negative qRT-PCR responses, track how effectively the patient's immune system is fighting the infection, and are potentially helpful for plasma transfusion therapies. In this work, based on the principle of localized surface plasmon resonance (LSPR), we develop an opto-microfluidic sensing platform with gold nanospikes, fabricated by electrodeposition, to detect the presence and amount of antibodies specific to the SARS-CoV-2 spike protein in 1µL of human plasma diluted in 1mL of buffer solution, within ∼30min. The target antibody concentration can be correlated with the LSPR wavelength peak shift of gold nanospikes caused by the local refractive index change due to the antigen-antibody binding. This label-free microfluidic platform achieves a limit of detection of ∼0.08ng/mL (∼0.5pM), falling under the clinical relevant concentration range. We demonstrate that our opto-microfluidic platform offers a promising point-of-care testing tool to complement standard serological assays and make SARS-CoV-2 quantitative diagnostics easier, cheaper, and faster.


Assuntos
Anticorpos Antivirais/sangue , Betacoronavirus/imunologia , Infecções por Coronavirus/sangue , Nanoestruturas/química , Pneumonia Viral/sangue , Glicoproteína da Espícula de Coronavírus/imunologia , Ressonância de Plasmônio de Superfície/instrumentação , Anticorpos Antivirais/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Desenho de Equipamento , Ouro/química , Humanos , Dispositivos Lab-On-A-Chip , Limite de Detecção , Nanoestruturas/ultraestrutura , Pandemias , Pneumonia Viral/imunologia , Pneumonia Viral/virologia
2.
Nat Commun ; 11(1): 4772, 2020 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-32973145

RESUMO

Surface-enhanced Raman spectroscopy (SERS) is a powerful tool for vibrational spectroscopy as it provides several orders of magnitude higher sensitivity than inherently weak spontaneous Raman scattering by exciting localized surface plasmon resonance (LSPR) on metal substrates. However, SERS can be unreliable for biomedical use since it sacrifices reproducibility, uniformity, biocompatibility, and durability due to its strong dependence on "hot spots", large photothermal heat generation, and easy oxidization. Here, we demonstrate the design, fabrication, and use of a metal-free (i.e., LSPR-free), topologically tailored nanostructure composed of porous carbon nanowires in an array as a SERS substrate to overcome all these problems. Specifically, it offers not only high signal enhancement (~106) due to its strong broadband charge-transfer resonance, but also extraordinarily high reproducibility due to the absence of hot spots, high durability due to no oxidization, and high compatibility to biomolecules due to its fluorescence quenching capability.


Assuntos
Carbono/química , Nanofios/química , Análise Espectral Raman/métodos , Fluorescência , Porosidade , Reprodutibilidade dos Testes , Ressonância de Plasmônio de Superfície/métodos , Propriedades de Superfície
3.
Biosens Bioelectron ; 166: 112436, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32750677

RESUMO

Our recent experience of the COVID-19 pandemic has highlighted the importance of easy-to-use, quick, cheap, sensitive and selective detection of virus pathogens for the efficient monitoring and treatment of virus diseases. Early detection of viruses provides essential information about possible efficient and targeted treatments, prolongs the therapeutic window and hence reduces morbidity. Graphene is a lightweight, chemically stable and conductive material that can be successfully utilized for the detection of various virus strains. The sensitivity and selectivity of graphene can be enhanced by its functionalization or combination with other materials. Introducing suitable functional groups and/or counterparts in the hybrid structure enables tuning of the optical and electrical properties, which is particularly attractive for rapid and easy-to-use virus detection. In this review, we cover all the different types of graphene-based sensors available for virus detection, including, e.g., photoluminescence and colorimetric sensors, and surface plasmon resonance biosensors. Various strategies of electrochemical detection of viruses based on, e.g., DNA hybridization or antigen-antibody interactions, are also discussed. We summarize the current state-of-the-art applications of graphene-based systems for sensing a variety of viruses, e.g., SARS-CoV-2, influenza, dengue fever, hepatitis C virus, HIV, rotavirus and Zika virus. General principles, mechanisms of action, advantages and drawbacks are presented to provide useful information for the further development and construction of advanced virus biosensors. We highlight that the unique and tunable physicochemical properties of graphene-based nanomaterials make them ideal candidates for engineering and miniaturization of biosensors.


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas Biossensoriais , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Grafite , Pneumonia Viral/diagnóstico , Vírus/isolamento & purificação , Reações Antígeno-Anticorpo , Betacoronavirus/genética , Betacoronavirus/patogenicidade , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Técnicas Biossensoriais/tendências , Técnicas de Laboratório Clínico/instrumentação , Técnicas de Laboratório Clínico/métodos , Técnicas de Laboratório Clínico/estatística & dados numéricos , Colorimetria , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , DNA Viral/análise , DNA Viral/genética , Técnicas Eletroquímicas , Desenho de Equipamento , Grafite/química , Humanos , Luminescência , Nanoestruturas/química , Hibridização de Ácido Nucleico , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , Pontos Quânticos/química , Análise Espectral Raman , Ressonância de Plasmônio de Superfície , Virologia/métodos , Vírus/genética , Vírus/patogenicidade
4.
Anal Chem ; 92(17): 11520-11524, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32786452

RESUMO

The traditional approach for analyzing interaction data from biosensors instruments is based on the simplified assumption that also larger biomolecules interactions are homogeneous. It was recently reported that the human receptor angiotensin-converting enzyme 2 (ACE2) plays a key role for capturing SARS-CoV-2 into the human target body, and binding studies were performed using biosensors techniques based on surface plasmon resonance and bio-layer interferometry. The published affinity constants for the interactions, derived using the traditional approach, described a single interaction between ACE2 and the SARS-CoV-2 receptor binding domain (RBD). We reanalyzed these data sets using our advanced four-step approach based on an adaptive interaction distribution algorithm (AIDA) that accounts for the great complexity of larger biomolecules and gives a two-dimensional distribution of association and dissociation rate constants. Our results showed that in both cases the standard assumption about a single interaction was erroneous, and in one of the cases, the value of the affinity constant KD differed more than 300% between the reported value and our calculation. This information can prove very useful in providing mechanistic information and insights about the mechanism of interactions between ACE2 and SARS-CoV-2 RBD or similar systems.


Assuntos
Betacoronavirus/química , Interferometria/estatística & dados numéricos , Peptidil Dipeptidase A/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Ressonância de Plasmônio de Superfície/estatística & dados numéricos , Algoritmos , Humanos , Cinética , Ligantes , Peptidil Dipeptidase A/química , Ligação Proteica , Domínios Proteicos , Glicoproteína da Espícula de Coronavírus/química
5.
PLoS One ; 15(7): e0236710, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32735606

RESUMO

Argonaute (AGO) proteins are the key component of the RNA interference machinery that suppresses gene expression by forming an RNA-induced silencing complex (RISC) with microRNAs (miRNAs). Each miRNA is involved in various cellular processes, such as development, differentiation, tumorigenesis, and viral infection. Thus, molecules that regulate miRNA function are expected to have therapeutic potential. In addition, the biogenesis of miRNA is a multistep process involving various proteins, although the complete pathway remains to be elucidated. Therefore, identification of molecules that can specifically modulate each step will help understand the mechanism of gene suppression. To date, several AGO2 inhibitors have been identified. However, these molecules were identified through a single screening method, and no studies have specifically evaluated a combinatorial strategy. Here, we demonstrated a combinatorial screening (SCR) approach comprising an in silico molecular docking study, surface plasmon resonance (SPR) analysis, and nuclear magnetic resonance (NMR) analysis, focusing on the strong binding between the 5'-terminal phosphate of RNA and the AGO2 middle (MID) domain. By combining SPR and NMR, we identified binding modes of amino acid residues binding to AGO2. First, using a large chemical library (over 6,000,000 compounds), 171 compounds with acidic functional groups were screened using in silico SCR. Next, we constructed an SPR inhibition system that could analyze only the 5'-terminal binding site of RNA, and nine molecules that strongly bound to the AGO2 MID domain were selected. Finally, using NMR, three molecules that bound to the desired site were identified. The RISC inhibitory ability of the "hit" compounds was analyzed in human cell lysate, and all three hit compounds strongly inhibited the binding between double-stranded RNA and AGO2.


Assuntos
Proteínas Argonauta/antagonistas & inibidores , Avaliação Pré-Clínica de Medicamentos/métodos , Espectroscopia de Ressonância Magnética , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Ressonância de Plasmônio de Superfície , Proteínas Argonauta/química , Proteínas Argonauta/metabolismo , Simulação por Computador , Células HeLa , Humanos , Simulação de Acoplamento Molecular , Conformação Proteica , Bibliotecas de Moléculas Pequenas/metabolismo
6.
PLoS One ; 15(8): e0236839, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32780746

RESUMO

The majority of chronic myeloid leukemia (CML) cases are caused by a chromosomal translocation linking the breakpoint cluster region (BCR) gene to the Abelson murine leukemia viral oncogene-1 (ABL1), creating the mutant fusion protein BCR-ABL1. Downstream of BCR-ABL1 is growth factor receptor-bound protein-2 (GRB2), an intracellular adapter protein that binds to BCR-ABL1 via its src-homology-2 (SH2) domain. This binding constitutively activates growth pathways, downregulates apoptosis, and leads to an over proliferation of immature and dysfunctional myeloid cells. Utilizing novel synthetic methods, we developed four furo-quinoxaline compounds as GRB2 SH2 domain antagonists with the goal of disrupting this leukemogenic signaling. One of the four antagonists, NHD2-15, showed a significant reduction in proliferation of K562 cells, a human BCR-ABL1+ leukemic cell line. To elucidate the mode of action of these compounds, various biophysical, in vitro, and in vivo assays were performed. Surface plasmon resonance (SPR) assays indicated that NHD2-15 antagonized GRB2, binding with a KD value of 119 ± 2 µM. Cellulose nitrate (CN) assays indicated that the compound selectively bound the SH2 domain of GRB2. Western blot assays suggested the antagonist downregulated proteins involved in leukemic transformation. Finally, NHD2-15 was nontoxic to primary cells and adult zebrafish, indicating that it may be an effective clinical treatment for CML.


Assuntos
Proliferação de Células/efeitos dos fármacos , Proteína Adaptadora GRB2/antagonistas & inibidores , Quinoxalinas/farmacologia , Animais , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Proteínas de Fusão bcr-abl/metabolismo , Proteína Adaptadora GRB2/química , Proteína Adaptadora GRB2/metabolismo , Humanos , Células K562 , Rim/citologia , Cinética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Ligação Proteica , Quinoxalinas/química , Quinoxalinas/metabolismo , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Ressonância de Plasmônio de Superfície , Peixe-Zebra , Domínios de Homologia de src
7.
ACS Sens ; 5(9): 2663-2678, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32786383

RESUMO

The global sanitary crisis caused by the emergence of the respiratory virus SARS-CoV-2 and the COVID-19 outbreak has revealed the urgent need for rapid, accurate, and affordable diagnostic tests to broadly and massively monitor the population in order to properly manage and control the spread of the pandemic. Current diagnostic techniques essentially rely on polymerase chain reaction (PCR) tests, which provide the required sensitivity and specificity. However, its relatively long time-to-result, including sample transport to a specialized laboratory, delays massive detection. Rapid lateral flow tests (both antigen and serological tests) are a remarkable alternative for rapid point-of-care diagnostics, but they exhibit critical limitations as they do not always achieve the required sensitivity for reliable diagnostics and surveillance. Next-generation diagnostic tools capable of overcoming all the above limitations are in demand, and optical biosensors are an excellent option to surpass such critical issues. Label-free nanophotonic biosensors offer high sensitivity and operational robustness with an enormous potential for integration in compact autonomous devices to be delivered out-of-the-lab at the point-of-care (POC). Taking the current COVID-19 pandemic as a critical case scenario, we provide an overview of the diagnostic techniques for respiratory viruses and analyze how nanophotonic biosensors can contribute to improving such diagnostics. We review the ongoing published work using this biosensor technology for intact virus detection, nucleic acid detection or serological tests, and the key factors for bringing nanophotonic POC biosensors to accurate and effective COVID-19 diagnosis on the short term.


Assuntos
Betacoronavirus , Infecções por Coronavirus/diagnóstico , Nanoestruturas/química , Pneumonia Viral/diagnóstico , Ressonância de Plasmônio de Superfície/métodos , Antígenos Virais/análise , Betacoronavirus/química , Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico , Genoma Viral , Humanos , Imunoensaio/métodos , Nanoestruturas/efeitos da radiação , Pandemias , Testes Sorológicos/métodos
8.
Sensors (Basel) ; 20(17)2020 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-32842601

RESUMO

The global burden of coronavirus disease 2019 (COVID-19) to public health and global economy has stressed the need for rapid and simple diagnostic methods. From this perspective, plasmonic-based biosensing can manage the threat of infectious diseases by providing timely virus monitoring. In recent years, many plasmonics' platforms have embraced the challenge of offering on-site strategies to complement traditional diagnostic methods relying on the polymerase chain reaction (PCR) and enzyme-linked immunosorbent assays (ELISA). This review compiled recent progress on the development of novel plasmonic sensing schemes for the effective control of virus-related diseases. A special focus was set on the utilization of plasmonic nanostructures in combination with other detection formats involving colorimetric, fluorescence, luminescence, or Raman scattering enhancement. The quantification of different viruses (e.g., hepatitis virus, influenza virus, norovirus, dengue virus, Ebola virus, Zika virus) with particular attention to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) was reviewed from the perspective of the biomarker and the biological receptor immobilized on the sensor chip. Technological limitations including selectivity, stability, and monitoring in biological matrices were also reviewed for different plasmonic-sensing approaches.


Assuntos
Betacoronavirus/isolamento & purificação , Técnicas Biossensoriais , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Ressonância de Plasmônio de Superfície/métodos , Betacoronavirus/patogenicidade , Colorimetria , Infecções por Coronavirus/virologia , Fluorescência , Humanos , Nanoestruturas/química , Pandemias , Pneumonia Viral/virologia , Análise Espectral Raman
9.
Opt Express ; 28(12): 18479-18492, 2020 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-32680046

RESUMO

Biomolecule sensing plays an important role in both fundamental biological studies and medical diagnostic applications. Infrared (IR) spectroscopy presents opportunities for sensing biomolecules as it allows their fingerprints to be determined by directly measuring their absorption spectra. However, the detection of biomolecules at low concentrations is difficult with conventional IR spectroscopy due to signal-to-noise considerations. This has led to recent interest on the use of nanostructured surfaces to boost the signals from biomolecules in a method termed surface enhanced infrared spectroscopy. So far, efforts have largely involved the use of metallic nanoantennas (which produce large field enhancement) or graphene nanostructures (which produce strong field confinement and provide electrical tunability). Here, we propose a nanostructured surface that combines the large field enhancement of metallic nanoantennas with the strong field confinement and electrical tunability of graphene plasmons. Our device consists of an array of plasmonic nanoantennas and graphene nanoslits on a resonant substrate. We perform systematic electromagnetic simulations to quantify the sensing performance of the proposed device and show that it outperforms designs in which only plasmons from metallic nanoantennas or plasmons from graphene are utilized. These investigations consider the model system of a representative protein-goat anti-mouse immunoglobulin G (IgG) - in monolayer or sub-monolayer form. Our findings provide guidance for future biosensors for the sensitive quantification and identification of biomolecules.


Assuntos
Grafite , Nanopartículas Metálicas , Espectrofotometria Infravermelho/instrumentação , Ressonância de Plasmônio de Superfície/instrumentação , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Desenho de Equipamento/métodos , Proteínas/análise , Ressonância de Plasmônio de Superfície/métodos
10.
Food Chem ; 332: 127431, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32645668

RESUMO

Illegal usage of ß-agonists as the animal growth promoters can lead to multiple harmful impacts to public health, thus detection of ß-agonists at trace level in complex sample matrixes is of great importance. In recent years, emergence of advanced nanomaterials greatly facilitates the advancement of sensors in terms of sensitivity, specificity and robustness. Plenty of nanoparticles-based sensors have been developed for ß-agonists determination. In this review, we comprehensively summarized the construction of emerging nanoparticles-based sensors (including colorimetric sensors, fluorescent sensors, chemiluminescent sensors, electrochemical sensors, electrochemiluminescent sensors, surface enhanced Raman scattering sensors, surface plasmon resonance sensors, quartz crystal microbalance sensors, etc.), and nanomaterial-based enzyme-linked immunosorbent assay (nano-ELISA). Impressively, the applications of nanoparticles-based sensors and nano-ELISAs in the detection of ß-agonists have also been summarized and discussed. In the end, future opportunities and challenges in the design construction of nanoparticles (NPs)-based sensors and their applications in ß-agonist assay are tentatively proposed.


Assuntos
Agonistas Adrenérgicos/análise , Nanoestruturas/química , Animais , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Colorimetria/instrumentação , Colorimetria/métodos , Humanos , Nanotecnologia/instrumentação , Nanotecnologia/métodos , Análise Espectral Raman/instrumentação , Análise Espectral Raman/métodos , Ressonância de Plasmônio de Superfície/instrumentação , Ressonância de Plasmônio de Superfície/métodos
11.
Antiviral Res ; 181: 104873, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32653452

RESUMO

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has resulted in a pandemic and continues to spread around the globe at an unprecedented rate. To date, no effective therapeutic is available to fight its associated disease, COVID-19. Our discovery of a novel insertion of glycosaminoglycan (GAG)-binding motif at S1/S2 proteolytic cleavage site (681-686 (PRRARS)) and two other GAG-binding-like motifs within SARS-CoV-2 spike glycoprotein (SGP) led us to hypothesize that host cell surface GAGs may interact SARS-CoV-2 SGPs to facilitate host cell entry. Using a surface plasmon resonance direct binding assay, we found that both monomeric and trimeric SARS-CoV-2 SGP bind more tightly to immobilized heparin (KD = 40 pM and 73 pM, respectively) than the SARS-CoV and MERS-CoV SGPs (500 nM and 1 nM, respectively). In competitive binding studies, the IC50 of heparin, tri-sulfated non-anticoagulant heparan sulfate, and non-anticoagulant low molecular weight heparin against SARS-CoV-2 SGP binding to immobilized heparin were 0.056 µM, 0.12 µM, and 26.4 µM, respectively. Finally, unbiased computational ligand docking indicates that heparan sulfate interacts with the GAG-binding motif at the S1/S2 site on each monomer interface in the trimeric SARS-CoV-2 SGP, and at another site (453-459 (YRLFRKS)) when the receptor-binding domain is in an open conformation. The current study serves a foundation to further investigate biological roles of GAGs in SARS-CoV-2 pathogenesis. Furthermore, our findings may provide additional basis for further heparin-based interventions for COVID-19 patients exhibiting thrombotic complications.


Assuntos
Betacoronavirus/metabolismo , Infecções por Coronavirus/virologia , Heparina/metabolismo , Pandemias , Pneumonia Viral/virologia , Síndrome Respiratória Aguda Grave/virologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Sítios de Ligação , Humanos , Cinética , Simulação de Acoplamento Molecular , Ligação Proteica , Ressonância de Plasmônio de Superfície
12.
Food Chem ; 332: 127346, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32619938

RESUMO

Fiber optic surface plasmon resonance (SPR) sensor utilizing silver (Ag) and Ag-graphene oxide (GO) is designed and developed for the detection of adulteration of glucose and fructose in pure honey. The concentration range of the two adulterants in pure honey is varied from 4% to 20% with a step change of 4%. The experiments were performed with two different fiber optic probes viz. Probe 1 and Probe 2. Probe 1 is fabricated by coating 50 nm Ag film on unclad optical fiber portion and Probe 2 is fabricated by modifying Ag film with GO for sensitivity improvement. The study confirms that using GO modified fiber optic probe, the sensitivity is enhanced to 24% and 37% for glucose and fructose adulterated honey samples respectively. The technique presented in this study is easy, rapid, label free and shows high prospective for the detection of adulterants in pure honey.


Assuntos
Grafite/química , Mel/análise , Fibras Ópticas , Prata/química , Ressonância de Plasmônio de Superfície/instrumentação
13.
PLoS One ; 15(6): e0233443, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32497056

RESUMO

Large (> 1 µm) tumor-derived extracellular vesicles (tdEVs) enriched from the cell fraction of centrifuged whole blood are prognostic in metastatic castration-resistant prostate cancer (mCRPC) patients. However, the highest concentration of tdEVs is expected in the cell-free plasma fraction. In this pilot study, we determine whether mCRPC patients can be discriminated from healthy controls based on detection of tdEVs (< 1µm, EpCAM+) and/or other EVs, in cell-free plasma and/or urine. The presence of marker+ EVs in plasma and urine samples from mCRPC patients (n = 5) and healthy controls (n = 5) was determined by flow cytometry (FCM) and surface plasmon resonance imaging (SPRi) using an antibody panel and lactadherin. For FCM, the concentrations of marker positive (+) particles and EVs (refractive index <1.42) were determined. Only the lactadherin+ particle and EV concentration in plasma measured by FCM differed significantly between patients and controls (p = 0.017). All other markers did not result in signals exceeding the background on both FCM and SPRi, or did not differ significantly between patients and controls. In conclusion, no difference was found between patients and controls based on the detection of tdEVs. For FCM, the measured sample volumes are too small to detect tdEVs. For SPRi, the concentration of tdEVs is probably too low to be detected. Thus, to detect tdEVs in cell-free plasma and/or urine, EV enrichment and/or concentration is required. Furthermore, we recommend testing other markers and/or a combination of markers to discriminate mCRPC patients from healthy controls.


Assuntos
Adenocarcinoma/secundário , Vesículas Extracelulares/metabolismo , Citometria de Fluxo/métodos , Neoplasias de Próstata Resistentes à Castração/sangue , Neoplasias de Próstata Resistentes à Castração/urina , Ressonância de Plasmônio de Superfície/métodos , Adenocarcinoma/sangue , Adenocarcinoma/urina , Idoso , Idoso de 80 Anos ou mais , Antígenos de Superfície/sangue , Biomarcadores Tumorais , Linhagem Celular Tumoral , Meios de Cultivo Condicionados , Vesículas Extracelulares/química , Humanos , Masculino , Proteínas do Leite/sangue , Proteínas de Neoplasias/sangue , Proteínas de Neoplasias/urina , Projetos Piloto , Neoplasias de Próstata Resistentes à Castração/patologia
14.
Nat Commun ; 11(1): 2908, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32518267

RESUMO

Adoptive cell therapy (ACT) with tumor-specific T cells can mediate cancer regression. The main target of tumor-specific T cells are neoantigens arising from mutations in self-proteins. Although the majority of cancer neoantigens are unique to each patient, and therefore not broadly useful for ACT, some are shared. We studied oligoclonal T-cell receptors (TCRs) that recognize a shared neoepitope arising from a driver mutation in the p53 oncogene (p53R175H) presented by HLA-A2. Here we report structures of wild-type and mutant p53-HLA-A2 ligands, as well as structures of three tumor-specific TCRs bound to p53R175H-HLA-A2. These structures reveal how a driver mutation in p53 rendered a self-peptide visible to T cells. The TCRs employ structurally distinct strategies that are highly focused on the mutation to discriminate between mutant and wild-type p53. The TCR-p53R175H-HLA-A2 complexes provide a framework for designing TCRs to improve potency for ACT without sacrificing specificity.


Assuntos
Antígenos de Neoplasias/química , Antígeno HLA-A2/química , Mutação , Linfócitos T/imunologia , Proteína Supressora de Tumor p53/química , Sítios de Ligação , Biotinilação , Códon , Cristalografia por Raios X , Epitopos , Escherichia coli/metabolismo , Humanos , Imunoterapia Adotiva , Ligantes , Linfócitos do Interstício Tumoral/imunologia , Neoplasias/metabolismo , Peptídeos/química , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Receptores de Antígenos de Linfócitos T/metabolismo , Software , Ressonância de Plasmônio de Superfície
15.
Nat Commun ; 11(1): 2930, 2020 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-32523000

RESUMO

Enhancement of optical emission on plasmonic nanostructures is intrinsically limited by the distance between the emitter and nanostructure surface, owing to a tightly-confined and exponentially-decaying electromagnetic field. This fundamental limitation prevents efficient application of plasmonic fluorescence enhancement for diversely-sized molecular assemblies. We demonstrate a three-dimensionally-tapered gap plasmon nanocavity that overcomes this fundamental limitation through near-homogeneous yet powerful volumetric confinement of electromagnetic field inside an open-access nanotip. The 3D-tapered device provides fluorescence enhancement factors close to 2200 uniformly for various molecular assemblies ranging from few angstroms to 20 nanometers in size. Furthermore, our nanostructure allows detection of low concentration (10 pM) biomarkers as well as specific capture of single antibody molecules at the nanocavity tip for high resolution molecular binding analysis. Overcoming molecule position-derived large variations in plasmonic enhancement can propel widespread application of this technique for sensitive detection and analysis of complex molecular assemblies at or near single molecule resolution.


Assuntos
Técnicas Biossensoriais , Ouro/química , Nanopartículas Metálicas/química , Nanoestruturas/química , Nanotecnologia/métodos , Ressonância de Plasmônio de Superfície
16.
Food Chem ; 329: 127160, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32485646

RESUMO

In this study, we developed an indirect competitive plasmonic immunoassay using glucose oxidase (GOx)-induced redox reaction to etch Au nanorods (AuNRs) for qualitative and quantitative detection of aflatoxin M1 (AFM1) in milk. In this system, streptavidin (SA) was selected as a linker between biotinylated anti-AFM1-monoantibody (bio-mAb) and biotinylated GOx (bio-GOx) to form the immunocomplexes bio-mAb-SA-bio-GOx. After the oxidation of the glucose and I-, the resultant I2 could etch cetytrimethylammonium bromide (CTAB)-stabilized AuNRs. This resulted in the blue shift of the longitudinal localized surface plasmon resonance (LSPR) extinction peak, with a color change from blue to pink. The linear range and limit of detection (LOD) of the plasmonic immunoassay were 0.25-10 ng mL-1 and 0.11 ng mL-1, respectively. It displayed quantitative detection with high sensitivity, specificity, recovery, and accuracy, which is promising for qualitative and quantitative detection of AFM1 in food safety.


Assuntos
Aflatoxina M1/análise , Contaminação de Alimentos/análise , Imunoensaio/métodos , Leite/química , Nanotubos , Animais , Glucose/química , Glucose Oxidase/química , Ouro/química , Imunoensaio/instrumentação , Limite de Detecção , Nanotubos/química , Sensibilidade e Especificidade , Ressonância de Plasmônio de Superfície/métodos
17.
Food Chem ; 331: 127163, 2020 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-32593037

RESUMO

Herein, a surface plasmon resonance (SPR) enhanced DNA biosensor has been developed for real-time detection of donkey meat marker using biotinylated reporter and streptavidin functionalized gold nanostars (Stre@GNSs). Compared to the direct detection assay, this sandwich format for the enhancement of the signal, resulted in 6-folds orders increase in the sensitivity. Target DNA could be detected with the lower limit of quantification (LLOQ) of 1.0 nM with a relative standard deviation (RSD, n = 3) of 0.85%. In addition, the fabricated SPR sensor showed good selectivity for the target analyte over full complementary, single-base mismatch, three base-mismatch and non-complementary oligonucleotides. Finally, the proposed sensor was successfully applied for detection of donkey meat adulteration with various percentages in homemade beef sausage, as a real sample. The results indicated that the proposed biosensor provides a high specificity, easy, good sensitivity and fast approach for identification of donkey meat adulteration in food samples.


Assuntos
Técnicas Biossensoriais/métodos , Equidae/genética , Contaminação de Alimentos/análise , Carne/análise , Animais , Técnicas Biossensoriais/instrumentação , DNA/análise , Análise de Alimentos/instrumentação , Análise de Alimentos/métodos , Ouro/química , Limite de Detecção , Microscopia Eletrônica de Transmissão , Nanoestruturas/química , Espectrofotometria Ultravioleta , Estreptavidina/química , Ressonância de Plasmônio de Superfície/métodos
18.
ACS Nano ; 14(6): 7617-7627, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32437124

RESUMO

The current outbreak of the pandemic coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) demands its rapid, convenient, and large-scale diagnosis to downregulate its spread within as well as across the communities. But the reliability, reproducibility, and selectivity of majority of such diagnostic tests fail when they are tested either to a viral load at its early representation or to a viral gene mutated during its current spread. In this regard, a selective "naked-eye" detection of SARS-CoV-2 is highly desirable, which can be tested without accessing any advanced instrumental techniques. We herein report the development of a colorimetric assay based on gold nanoparticles (AuNPs), when capped with suitably designed thiol-modified antisense oligonucleotides (ASOs) specific for N-gene (nucleocapsid phosphoprotein) of SARS-CoV-2, could be used for diagnosing positive COVID-19 cases within 10 min from the isolated RNA samples. The thiol-modified ASO-capped AuNPs agglomerate selectively in the presence of its target RNA sequence of SARS-CoV-2 and demonstrate a change in its surface plasmon resonance. Further, the addition of RNaseH cleaves the RNA strand from the RNA-DNA hybrid leading to a visually detectable precipitate from the solution mediated by the additional agglomeration among the AuNPs. The selectivity of the assay has been monitored in the presence of MERS-CoV viral RNA with a limit of detection of 0.18 ng/µL of RNA having SARS-CoV-2 viral load. Thus, the current study reports a selective and visual "naked-eye" detection of COVID-19 causative virus, SARS-CoV-2, without the requirement of any sophisticated instrumental techniques.


Assuntos
Betacoronavirus/genética , Técnicas Biossensoriais/métodos , Infecções por Coronavirus/diagnóstico , Nanopartículas Metálicas , Proteínas do Nucleocapsídeo/genética , Oligonucleotídeos Antissenso/genética , Pneumonia Viral/diagnóstico , Sequência de Bases , Betacoronavirus/isolamento & purificação , Colorimetria/métodos , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Genes Virais , Ouro , Humanos , Nanopartículas Metálicas/ultraestrutura , Microscopia Eletrônica de Transmissão , Nanotecnologia/métodos , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , Capuzes de RNA/genética , RNA Viral/genética , Ressonância de Plasmônio de Superfície/métodos
19.
Cancer Immunol Immunother ; 69(9): 1833-1840, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32350593

RESUMO

BACKGROUND: Bladder cancer is diagnosed by the use of several biomarkers, including survivin. This protein has an important role in the cancer progression by controlling the rate of cell apoptosis. Findings show that there is no survivin in normal tissues, whereas the level of survivin expression increases in tumor cells. DESIGN: The purpose of this study was to specify the reactive antibodies to survivin protein as a biomarker to determine the bladder cancer stage with ELISA method and using GNPs conjugated with survivin antibody. The serum and urine samples of patients with bladder cancer were collected among those referred to Sina Hospital, Tehran, Iran. The survivin protein level was measured in the serum and urine by ELISA technique and in the urine by GNPs conjugated with survivin. RESULTS: Based on the results of ELISA, the serum and urinary levels of survivin increased significantly in T3 and T4 stages of the disease (high grades), compared with the healthy individuals. Also, using conjugated GNPs, survivin protein was detected in the urine specimens of patients at all grades (low and high grades). CONCLUSION: Our findings showed that using the ELISA technique, the increased level of survivin could be identified in high grades of bladder cancer, but using anti-survivin antibody-conjugated GNPs, bladder cancer can be detected in early stages. The applied method was found to be a rapid tool, dependent on visible color changes and colorimetric detection, without any need for reader devices.


Assuntos
Anticorpos/metabolismo , Ouro/administração & dosagem , Nanopartículas Metálicas/administração & dosagem , Survivina/metabolismo , Neoplasias da Bexiga Urinária/diagnóstico , Neoplasias da Bexiga Urinária/metabolismo , Biomarcadores Tumorais/metabolismo , Feminino , Humanos , Masculino , Sensibilidade e Especificidade , Ressonância de Plasmônio de Superfície/métodos
20.
Ecotoxicol Environ Saf ; 200: 110771, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32464443

RESUMO

Herein is presented a simple and sensible method to determine organic pollutants in water, based on the utilization of silver nanoparticles (AgNPs) loaded in Polyacrylamide (PAAm)/starch hybrid hydrogels combined with surface-enhanced Raman scattering (SERS) spectroscopy. The materials were characterized by swelling degree studies, UV-Visible spectroscopy (UV-Vis), X-ray diffraction (XRD) and scanning electron microscopy (SEM). PAAm/starch hydrogels showed variable swelling capacity, according to the synthetic molar composition. The most promising results were attributed to lower concentrations of starch and crosslink agent (N,N'-methylenebisacrylamide - MBA). Spectroscopic analysis confirmed the formation of AgNPs, by noticing the peak at around 420 nm, due to its surface plasmon resonance (SPR) effect. The results showed that AgNPs were stabilized by hydrogels networks. The average size of the AgNPs was smaller than 100 nm and the size and quantity of nanoparticles were influenced by the molar composition of the hydrogel matrix. The SERS substrate based on the AgNPs-PAAm/starch exhibited reproducibility, stability, and limit of detection (LOD) of phenol in water of 1 × 10-8 M. The average mass of AgNPs-PAAm/starch hydrogels used for each detection analysis was around 10 mg. The spectra with enhanced intensities were possible due to a large number of hot spots generated on the AgNPs-PAAm/starch hydrogel substrate, which leads to potential use for organic pollutant detection. In addition, there is also the possibility of reusing the hydrogel matrix substrate in other analyzes.


Assuntos
Monitoramento Ambiental/métodos , Hidrogéis/química , Nanopartículas Metálicas/química , Fenóis/análise , Prata/química , Poluentes Químicos da Água/análise , Resinas Acrílicas/química , Monitoramento Ambiental/instrumentação , Limite de Detecção , Microscopia Eletrônica de Varredura , Reprodutibilidade dos Testes , Análise Espectral Raman , Amido/química , Ressonância de Plasmônio de Superfície , Água , Difração de Raios X
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