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1.
ACS Sens ; 5(1): 13-18, 2020 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-31833351

RESUMO

Environmental hazards typically are encountered in the gaseous phase; however, selective sensing modalities for identifying and quantitating compounds of interest in an inexpensive, pseudo-real-time format are severely lacking. Here, we present a novel proof-of-concept that combines an Air2Liquid sampler in conjunction with an oil-in-water microfluidic assay for detection of organophosphates. We believe this proof-of-concept will enable development of a new platform technology for semivolatile detection that we have demonstrated to detect 50 pmoles (2 ppb) of neurotoxic organophosphates.


Assuntos
Técnicas Biossensoriais/métodos , Gases/química , Organofosfatos/metabolismo
2.
Contemp Clin Trials ; 68: 7-13, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29535047

RESUMO

INTRODUCTION: Pain is one of the most burdensome symptoms associated with cancer and its treatment, and opioids are the cornerstone of pain management. Opioid therapy is empirically selected, and patients often require adjustments in therapy to effectively alleviate pain or ameliorate adverse drug effects that interfere with quality of life. There are data suggesting CYP2D6 genotype may contribute to inter-patient variability in response to opioids through its effects on opioid metabolism. Therefore, we aim to determine if CYP2D6 genotype-guided opioid prescribing results in greater reductions in pain and symptom severity and interference with daily living compared to a conventional prescribing approach in patients with cancer. METHODS: Patients with solid tumors with metastasis and a self-reported pain score ≥ 4/10 are eligible for enrollment and randomized to a genotype-guided or conventional pain management strategy. For patients in the genotype-guided arm, CYP2D6 genotype information is integrated into opioid prescribing decisions. Patients are asked to complete questionnaires regarding their pain, symptoms, and quality of life at baseline and 2, 4, 6, and 8 weeks after enrollment. The primary endpoint is differential change in pain severity by treatment strategy (genotype-guided versus conventional pain management). Secondary endpoints include change in pain and symptom interference with daily living. CONCLUSION: Pharmacogenetic-guided opioid selection for cancer pain management has potential clinical utility, but current evidence is limited to retrospective and observational studies. Precision Medicine Guided Treatment for Cancer Pain is a pragmatic clinical trial that seeks to determine the utility of CYP2D6 genotype-guided opioid prescribing in patients with cancer.


Assuntos
Analgésicos Opioides/uso terapêutico , Dor do Câncer , Neoplasias/complicações , Medição da Dor/métodos , Dor do Câncer/diagnóstico , Dor do Câncer/tratamento farmacológico , Dor do Câncer/genética , Citocromo P-450 CYP2D6/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Estadiamento de Neoplasias , Neoplasias/patologia , Manejo da Dor/métodos , Seleção de Pacientes , Farmacogenética/métodos , Ensaios Clínicos Pragmáticos como Assunto , Medicina de Precisão/métodos , Índice de Gravidade de Doença
3.
Langmuir ; 34(5): 2139-2146, 2018 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-29283584

RESUMO

DNA aptamers that bind biomolecular targets are of interest as the recognition element in colorimetric sensors based on gold nanoparticles (AuNP), where sensor functionality is related to changes in AuNP colloidal stability upon target binding. In order to understand the role of target binding on DNA-AuNP colloidal stability, we have used high-resolution NMR to characterize the interactions of the 36 nucleotide cocaine-binding aptamer (MN4) and related aptamers with AuNPs, cocaine, and cocaine metabolites. Changes in the aptamer imino proton NMR spectra with low (20 nM) concentrations of AuNP show that the aptamers undergo fast-exchange adsorption on the nanoparticle surface. An analysis of the spectral changes and the comparison with modified MN4 aptamers shows that the AuNP binding domain is localized on stem two of the three-stemmed aptamer. The identification of an AuNP recognition domain allows for the incorporation of AuNP binding functionality into a wide variety of aptamers. AuNP-induced spectral changes are not observed for the aptamer-AuNP mixtures in the presence of cocaine, demonstrating that aptamer absorption on the AuNP surface is modulated by aptamer-target interactions. The data also show that the DNA-AuNP interactions and sensor functionality are critically dependent on aptamer folding.


Assuntos
Aptâmeros de Nucleotídeos/química , Ouro/química , Nanopartículas Metálicas/química , Aptâmeros de Nucleotídeos/genética , Sequência de Bases , Concentração Osmolar , Temperatura
4.
J Vis Exp ; (96): e52545, 2015 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-25870978

RESUMO

Small molecules provide rich targets for biosensing applications due to their physiological implications as biomarkers of various aspects of human health and performance. Nucleic acid aptamers have been increasingly applied as recognition elements on biosensor platforms, but selecting aptamers toward small molecule targets requires special design considerations. This work describes modification and critical steps of a method designed to select structure-switching aptamers to small molecule targets. Binding sequences from a DNA library hybridized to complementary DNA capture probes on magnetic beads are separated from nonbinders via a target-induced change in conformation. This method is advantageous because sequences binding the support matrix (beads) will not be further amplified, and it does not require immobilization of the target molecule. However, the melting temperature of the capture probe and library is kept at or slightly above RT, such that sequences that dehybridize based on thermodynamics will also be present in the supernatant solution. This effectively limits the partitioning efficiency (ability to separate target binding sequences from nonbinders), and therefore many selection rounds will be required to remove background sequences. The reported method differs from previous structure-switching aptamer selections due to implementation of negative selection steps, simplified enrichment monitoring, and extension of the length of the capture probe following selection enrichment to provide enhanced stringency. The selected structure-switching aptamers are advantageous in a gold nanoparticle assay platform that reports the presence of a target molecule by the conformational change of the aptamer. The gold nanoparticle assay was applied because it provides a simple, rapid colorimetric readout that is beneficial in a clinical or deployed environment. Design and optimization considerations are presented for the assay as proof-of-principle work in buffer to provide a foundation for further extension of the work toward small molecule biosensing in physiological fluids.


Assuntos
Técnicas Biossensoriais/métodos , Colorimetria/métodos , Ouro/química , Nanopartículas Metálicas/química , Técnica de Seleção de Aptâmeros/métodos , Aptâmeros de Nucleotídeos/química , DNA Complementar/química , Humanos , Hidrocortisona/análise , Hidrocortisona/sangue
5.
Talanta ; 121: 247-55, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24607135

RESUMO

The feasibility of using aptamer-gold nanoparticle conjugates (Apt-AuNPs) to design colorimetric assays for in the field detection of small molecules was investigated. An assay to detect cocaine was designed using two clones of a known cocaine-binding aptamer. The assay was based on the AuNPs difference in affinity for single-stranded DNA (non-binding) and double stranded DNA (target bound). In the first assay, a commonly used design was followed, in which the aptamer and target were incubated to allow binding followed by exposure to the AuNPs. Interactions between the non-bound analytes and the AuNPs surface resulted in a number of false positives. The assay was redesigned by incubating the AuNPs and the aptamer prior to target addition to passivate the AuNPs surface. The adsorbed aptamer was able to bind the target while preventing non-specific interactions. The assay was validated with a number of masking and cutting agents and other controlled substances showing minimal false positives. Studies to improve the assay performance in the field were performed, showing that assay activity could be preserved for up to 2 months. To facilitate the assay analysis, an android application for automatic colorimetric characterization was developed. The application was validated by challenging the assay with cocaine standards of different concentrations, and comparing the results to a conventional plate reader, showing outstanding agreement. Finally, the rapid identification of cocaine in mixtures mimicking street samples was demonstrated. This work established that Apt-AuNPs can be used to design robust assays to be used in the field.

6.
Fitoterapia ; 93: 62-6, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24370663

RESUMO

Five anthranilic acid derivatives, a mixture I of three new compounds 11'-hexadecenoylanthranilic acid (1), 9'-hexadecenoylanthranilic acid (2), and 7'-hexadecenoylanthranilic acid (3), as well as a new compound 9,12,15-octadecatrienoylanthranilic acid (4) together with a new natural product, hexadecanoylanthranilic acid (5), were isolated from Geijera parviflora Lindl. (Rutaceae). Their structures were elucidated by extensive spectroscopic measurements, and the positions of the double bonds in compounds 1-3 of the mixture I were determined by tandem mass spectrometry employing ozone-induced dissociation. The mixture I and compound 5 showed good antibacterial activity against several Gram-positive strains.


Assuntos
Antibacterianos/isolamento & purificação , Rutaceae/química , ortoaminobenzoatos/química , Antibacterianos/química , Testes de Sensibilidade Microbiana
7.
J Nat Prod ; 76(7): 1384-7, 2013 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-23848189

RESUMO

Two novel alkaloids (parvifloranines A and B), possessing an unusual 11-carbon skeleton linked with amino acids, were isolated from Geijera parviflora, an endemic Australian Rutaceae. Their structures were elucidated by extensive spectroscopic measurements including 2D NMR analyses. Parvifloranine A was found to be a mixture of two enantiomers, (S)-1 and (R)-1, in a ratio of 1:4, based on their separation using a chiral column. Parvifloranine B is also believed to be a mixture of enantiomers. Proposed biosynthetic pathways are discussed. Parvifloranine A inhibited the synthesis of nitric oxide in LPS-stimulated RAW 264.7 macrophages with an IC50 value of 23.4 µM.


Assuntos
Alcaloides/isolamento & purificação , Alcaloides/química , Alcaloides/farmacologia , Animais , Austrália , Concentração Inibidora 50 , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Camundongos , Estrutura Molecular , Óxido Nítrico/biossíntese , Ressonância Magnética Nuclear Biomolecular , Estereoisomerismo
8.
Inorg Chem ; 52(11): 6311-8, 2013 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-23692439

RESUMO

Encapsulation of macrocyclic europium(III) chelates by discrete, monodisperse SiO2 nanoparticles (NPs) has been carried out, and the resulting significant enhancement of metal-derived luminescence has been studied to rationalize this dramatic effect. The tetraiminodiphenolate motif chosen for this study is easily synthesized and incorporated into the NP matrix under ambient conditions. The free complex exhibits primarily weak ligand-derived emission at room temperature, typical for these compounds, and displays intense metal-centered luminescence from the europium only when cooled to 77 K. Upon encapsulation by the NPs, however, europium-derived luminescence is visibly "turned on" at room temperature, yielding strong emission peaks characteristic of europium(III) with a corresponding enhancement factor of 6 × 10(6). The similar ligand singlet and triplet excited-state energies determined for the free complex (20820 and 17670 cm(-1), respectively) versus the encapsulated complex (20620 and 17730 cm(-1)) indicate that encapsulation does not affect the energy levels of the ligand appreciably. Instead, a detailed analysis of the metal-centered emission and ligand singlet and triplet emission bands for the free and encapsulated complexes reveals that the enhanced metal emission is due to the rigid environment afforded by the silica NP matrix affecting vibrationally mediated energy transfer. Further, the metal-centered emission lifetimes in methanol versus deuterated methanol indicate a decrease in the number of coordinated solvent molecules upon encapsulation, changing from an average of 3.3 to 2.1 bound methanol molecules and reducing the known quenching effect on europium-centered luminescence due to nearby OH vibrations.


Assuntos
Európio/química , Luminescência , Nanocápsulas/química , Compostos Organometálicos/química , Estrutura Molecular , Compostos Organometálicos/síntese química , Dióxido de Silício/química
9.
Anal Bioanal Chem ; 404(8): 2233-9, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22918539

RESUMO

The genotoxicity of a molecule refers to its ability to interact with DNA in a way that inhibits normal DNA replication and transcription possibly leading to mutagenesis or carcinogenesis. Assessing the genotoxicity of a compound is critical in the development of pharmaceuticals and other products designed for human consumption or use. Typically genotoxicity is established using expensive and time consuming methods using animals or bacteria like the Ames test, mouse lymphoma assay, or mouse and rat carcinogenicity tests. We have developed a magnetic nanoparticle-based assay that uses conjugated double-stranded DNA to serve as a substrate for interaction with genotoxic molecules. After application of a magnetic field, the genotoxic molecules are extracted with the DNA-conjugated magnetic nanoparticles. The genotoxic molecules can then be released and detected. To evaluate the potential of this assay, we have screened several genotoxic and non-genotoxic compounds and have demonstrated the ability to extract a genotoxic compound in the presence of a non-genotoxic molecule. The assay demonstrates suitable analytical performance and the ability to differentiate between genotoxic and non-genotoxic molecules providing a rapid and inexpensive alternative to more traditional methods of evaluating genotoxicity.


Assuntos
DNA/química , Magnetismo , Testes de Mutagenicidade/métodos , Mutagênicos/química , Nanopartículas/química , Cromatografia Líquida de Alta Pressão , Docetaxel , Doxorrubicina/química , Etídio/química , Indometacina/química , Dióxido de Silício/química , Propriedades de Superfície , Taxoides/química
10.
Anal Chem ; 83(3): 727-34, 2011 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-21218774

RESUMO

Aptamer-conjugated nanoparticles (ACNPs) have been used for a variety of applications, particularly dual nanoparticles for magnetic extraction and fluorescent labeling. In this type of assay, silica-coated magnetic and fluorophore-doped silica nanoparticles are conjugated to highly selective aptamers to detect and extract targeted cells in a variety of matrixes. However, considerable improvements are required in order to increase the selectivity and sensitivity of this two-particle assay to be useful in a clinical setting. To accomplish this, several parameters were investigated, including nanoparticle size, conjugation chemistry, use of multiple aptamer sequences on the nanoparticles, and use of multiple nanoparticles with different aptamer sequences. After identifying the best-performing elements, the improvements made to this assay's conditional parameters were combined to illustrate the overall enhanced sensitivity and selectivity of the two-particle assay using an innovative multiple aptamer approach, signifying a critical feature in the advancement of this technique.


Assuntos
Aptâmeros de Nucleotídeos/análise , Nanopartículas/química , Neoplasias/química , Aptâmeros de Nucleotídeos/química , Linhagem Celular Tumoral , Corantes Fluorescentes/análise , Corantes Fluorescentes/química , Humanos , Magnetismo , Tamanho da Partícula
11.
Anal Biochem ; 410(1): 124-32, 2011 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-21078282

RESUMO

Biosensors based on antibody recognition have a wide range of monitoring applications that apply to clinical, environmental, homeland security, and food problems. In an effort to improve the limit of detection of the Naval Research Laboratory (NRL) Array Biosensor, magnetic nanoparticles (MNPs) were designed and tested using a fluorescence-based array biosensor. The MNPs were coated with the fluorescently labeled protein, AlexaFluor647-chicken IgG (Alexa647-chick IgG). Antibody-labeled MNPs (Alexa647-chick-MNPs) were used to preconcentrate the target via magnetic separation and as the tracer to demonstrate binding to slides modified with anti-chicken IgG as a capture agent. A full optimization study of the antibody-modified MNPs and their use in the biosensor was performed. This investigation looked at the Alexa647-chick-MNP composition, MNP surface modifications, target preconcentration conditions, and the effect that magnetic extraction has on the Alexa647-chick-MNP binding with the array surface. The results demonstrate the impact of magnetic extraction using the MNPs labeled with fluorescent proteins both for target preconcentration and for subsequent integration into immunoassays performed under flow conditions for enhanced signal generation.


Assuntos
Métodos Analíticos de Preparação de Amostras/métodos , Técnicas Biossensoriais/métodos , Imunoensaio/métodos , Imunoglobulina G/química , Magnetismo , Nanopartículas/química , Animais , Técnicas Biossensoriais/instrumentação , Corantes Fluorescentes/química , Imunoensaio/instrumentação , Imunoglobulina G/imunologia , Limite de Detecção , Espectrometria de Fluorescência
12.
Anal Chem ; 80(4): 1067-72, 2008 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-18198894

RESUMO

Early and accurate detection of cancer often requires time-consuming techniques and expensive instrumentation. To address these limitations, we developed a colorimetric assay for the direct detection of diseased cells. The assay uses aptamer-conjugated gold nanoparticles to combine the selectivity and affinity of aptamers and the spectroscopic advantages of gold nanoparticles to allow for the sensitive detection of cancer cells. Samples with the target cells present exhibited a distinct color change while nontarget samples did not elicit any change in color. The assay also showed excellent sensitivity with both the naked eye and based on absorbance measurements. In addition, the assay was able to differentiate between different types of target and control cells based on the aptamer used in the assay indicating the wide applicability of the assay for diseased cell detection. On the basis of these qualities, aptamer-conjugated gold nanoparticles could become a powerful tool for point of care diagnostics.


Assuntos
Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Colorimetria/métodos , Ouro/química , Nanopartículas Metálicas/química , Neoplasias/patologia , Sequência de Bases , Microscopia Eletrônica de Transmissão/métodos , Neoplasias/ultraestrutura , Sensibilidade e Especificidade
13.
J Ethnopharmacol ; 112(2): 386-93, 2007 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-17485184

RESUMO

This paper reports on the isolation and identification of antibacterial constituents from the indigenous Australian medicinal plant Eremophila duttonii F. Muell. (Myoporaceae). Preparations derived from this plant are used by indigenous populations in the topical treatment of minor wounds, otitis and ocular complaints, and as a gargle for sore throat. Several authors have reported extracts of this plant to effect rapid bacteriolysis and inhibit growth of a wide range of Gram-positive micro-organisms. In other studies involving screening of native medicinal plants for antibacterial activity, extracts of Eremophila duttonii have been reported to consistently exhibit the highest potency amongst all species included. From a hexane extract, we identified two diterpenes of the serrulatane class, the principal constituents responsible for antibacterial activity and present as major constituents of the resinous leaf cuticle: serrulat-14-en-7,8,20-triol (1) and serrulat-14-en-3,7,8,20-tetraol (2). In addition, a hydroxylated furanosesquiterpene with mild antibacterial activity which appeared to be a novel compound was isolated from the extract and tentatively identified as 4-hydroxy-4-methyl-1-(2,3,4,5-tetrahydro-5-methyl[2,3'-bifuran]-5-yl) pentan-2-one. Minimum inhibitory concentrations for each of the compounds against three Gram-positive bacteria: Staphylococcus aureus (ATCC 29213), Staphylococcus epidermidis (ATCC 12228) and Streptococcus pneumoniae (ARL 10582), were determined using a micro-titre plate broth dilution assay.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Myoporaceae/química , Austrália , Bioensaio , Cromatografia em Camada Delgada , Hexanos , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Solventes , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus epidermidis/efeitos dos fármacos , Streptococcus pneumoniae/efeitos dos fármacos
14.
Anal Chem ; 79(8): 3075-82, 2007 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-17348633

RESUMO

We have extended the use the aptamer-conjugated nanoparticles for the collection and detection of multiple cancer cells. The aptamers were selected using a cell-based SELEX strategy in our laboratory for cancer cells that, when utilized in this method, allow for the selective recognition of the cells from complex mixtures including fetal bovine serum samples. Aptamer-conjugated magnetic nanoparticles were used for the selective targeting cell extraction, and aptamer-conjugated fluorescent nanoparticles were employed for sensitive cellular detection. Employing both types of nanoparticles allows for selective and sensitive detection not possible by using the particles separately. Fluorescent nanoparticles amplify the signal intensity versus a single fluorophore label resulting in improved sensitivity. In addition, aptamer-conjugated magnetic nanoparticles allow for extraction and enrichment of target cells not possible with other separation methods. Fluorescent imaging and a microplate reader were used for cellular detection to demonstrate the wide applicability of this methodology for medical diagnostics and cell enrichment and separation.


Assuntos
Aptâmeros de Nucleotídeos/química , Linfoma de Burkitt/patologia , Leucemia-Linfoma de Células T do Adulto/patologia , Linfoma de Células B/patologia , Nanopartículas/química , Técnica de Seleção de Aptâmeros/métodos , Linfoma de Burkitt/genética , Linhagem Celular Tumoral , Corantes Fluorescentes/química , Humanos , Leucemia-Linfoma de Células T do Adulto/genética , Linfoma de Células B/genética , Magnetismo
15.
Anal Chem ; 78(9): 2918-24, 2006 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-16642976

RESUMO

We have developed a method for the rapid collection and detection of leukemia cells using a novel two-nanoparticle assay with aptamers as the molecular recognition element. An aptamer sequence was selected using a cell-based SELEX strategy in our laboratory for CCRF-CEM acute leukemia cells that, when applied in this method, allows for specific recognition of the cells from complex mixtures including whole blood samples. Aptamer-modified magnetic nanoparticles were used for target cell extraction, while aptamer-modified fluorescent nanoparticles were simultaneously added for sensitive cell detection. Combining two types of nanoparticles allows for rapid, selective, and sensitive detection not possible by using either particle alone. Fluorescent nanoparticles amplify the signal intensity corresponding to a single aptamer binding event, resulting in improved sensitivity over methods using individual dye-labeled probes. In addition, aptamer-modified magnetic nanoparticles allow for rapid extraction of target cells not possible with other separation methods. Fluorescent imaging and flow cytometry were used for cellular detection to demonstrate the potential application of this method for medical diagnostics.


Assuntos
Aptâmeros de Nucleotídeos/química , Leucemia/diagnóstico , Nanopartículas/química , Linhagem Celular Tumoral , Separação Celular , Citometria de Fluxo/métodos , Corantes Fluorescentes/química , Humanos , Magnetismo , Técnica de Seleção de Aptâmeros/métodos , Sensibilidade e Especificidade
16.
Rapid Commun Mass Spectrom ; 18(20): 2367-74, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15386634

RESUMO

Nanoparticles for the extraction of peptides and subsequent analysis using atmospheric pressure matrix-assisted laser desorption/ionization (APMALDI) have been evaluated. The atmospheric pressure source allows for particles to be directly introduced in the liquid matrix, minimizing sample loss and analysis time. Described in this work are two sample preparation procedures for liquid APMALDI analysis: a C18 functionalized silica nanoparticle for hydrophobic extractions, and an aptamer functionalized magnetite core nanoparticle for rapid, affinity extractions. The C18 particles provide a non-selective support for rapid profiling applications, while the aptamer particles are directed towards reducing the complexity in biological samples. The aptamer functionalized particles provide a more selective analyte-nanoparticle interaction whereby the tertiary structure of the analyte becomes more critical to the extraction. In both cases, the liquid APMALDI matrix provides a support for ionization, and acts as the releasing agent for the analyte-particle interaction. Additionally, analyte enrichment was possible due to the large surface-to-volume ratio of the particles. The experiments conducted with functionalized nanoparticles, in an atmospheric pressure liquid matrix, present a basis for further methodologies and utilities of silica nanoparticles to be developed.


Assuntos
Fracionamento Químico/métodos , Cromatografia de Afinidade/métodos , Nanotubos/química , Peptídeos/análise , Peptídeos/química , Dióxido de Silício/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Adsorção , Sequência de Aminoácidos , Pressão Atmosférica , Cromatografia Líquida/métodos , Dados de Sequência Molecular , Nanotubos/ultraestrutura , Tamanho da Partícula , Ligação Proteica
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