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Human liver microsomes containing various drug-metabolizing cytochrome P450 (P450) enzymes, along with their NADPH-reductase bound to phospholipid membranes, were absorbed onto 1-pyrene butylamine pi-pi stacked with amine-functionalized multiwalled carbon nanotube-modified graphite electrodes. The interfaced microsomal biofilm demonstrated direct electrochemical communication with the underlying electrode surface and enhanced oxygen reduction electrocatalytic activity typical of heme enzymes such as P450s over the unmodified electrodes and nonenzymatic currents. Similar enhancements in currents were observed when the bioelectrodes were constructed with recombinant P450 2C9 (single isoform) expressed bactosomes. The designed liver microsomal and 2C9 bactosomal bioelectrodes successfully facilitated the electrocatalytic conversion of diclofenac, a drug candidate, into 4'-hydroxydiclofenac. The enzymatic electrocatalytic metabolite yield was several-fold greater on the modified electrodes than on the unmodified bulk graphite electrodes adsorbed with a microsomal or bactosomal film. The nonenzymatic metabolite production was less than the enzymatically catalyzed metabolite yield in the designed microsomal and bactosomal biofilm electrodes. To test the throughput potential of the designed biofilms, eight-electrode array configurations were tested with the microsomal and bactosomal biofilms toward electrochemical 4'-hydroxydiclofenac metabolite production from diclofenac. The stability of the designed microsomal bioelectrode was assessed using nonfaradaic impedance spectroscopy over 40 h, which indicated good stability.
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Diclofenaco , Diclofenaco/análogos & derivados , Grafite , Humanos , Diclofenaco/análise , Diclofenaco/metabolismo , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Sistema Enzimático do Citocromo P-450/análise , Sistema Enzimático do Citocromo P-450/metabolismo , EletrodosRESUMO
Here we describe the development of a dual electrochemical immunosensor microchip for simultaneous detection of insulin (I) and cortisol (C) biomarkers that can enhance the ability to improve glucose regulation using automated insulin delivery. The successful realization of the simultaneous I and C measurements has been realized by integrating different enzymatically-tagged competitive and sandwich immunoassay formats on a single chip platform. The insulin detection is based on a peroxidase (HRP)-labeled sandwich assay whereas the cortisol detection relies on an alkaline phosphatase (ALP)-labeled competitive immunoassay. The attractive analytical performance of the dual marker immunosensor, with no apparent cross-talk, was achieved through systematic optimization of the incubation and amperometric detection of the different captured enzyme tags. Evaluation of dual biosensor chip in untreated serum samples indicated favorable simultaneous detection of picomolar (pM) insulin and nanomolar (nM) cortisol concentrations in a single microliter sample droplet within less than 25min. The new dual immunosensor chip offers considerable promise for frequent decentralized testing of I and C towards a tighter glycemic control and improved management of diabetes.
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Técnicas Biossensoriais , Eletrodos , Hidrocortisona , Imunoensaio , InsulinaRESUMO
We report a large amplification of surface plasmon signals for a double hybridization microarray chip assembly that bridges localized gold and detection probe-carrying-core/shell Fe3O4@Au nanoparticles for detection of as low as 80 aM miRNA-155 marker in solution. The plasmonic wavelength match of the gold shell with surface localized gold nanoparticles and the additional scattering band of the core/shell material in resonance with the incident 800 nm light source are the underlying factors for the observed remarkable analyte signal at ultra-low (10-18 order) concentrations.
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Ouro , Nanopartículas Metálicas , Hibridização de Ácido Nucleico , Nucleotídeos , Ressonância de Plasmônio de SuperfícieRESUMO
Low-cost, voltage-driven biocatalytic designs for rapid drug metabolism assay, chemical toxicity screening, and pollutant biosensing represent considerable significance for pharmaceutical, biomedical, and environmental applications. In this study, we have designed biointerfaces of human liver microsomes with various roughened, high-purity graphite disk electrodes to study electrochemical and electrocatalytic properties. Successful spectral and microscopic characterizations, direct bioelectronic communication, direct electron-transfer rates from the electrode to liver microsomal enzymes, microsomal heme-enzyme specific oxygen reduction currents, and voltage-driven diclofenac hydroxylation (chosen as the probe reaction) are presented.
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Sistema Enzimático do Citocromo P-450/metabolismo , Técnicas Eletroquímicas , Grafite/metabolismo , Microssomos Hepáticos/metabolismo , Sistema Enzimático do Citocromo P-450/química , Eletrodos , Grafite/química , Humanos , Microssomos Hepáticos/química , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Electrode materials play an important role on the electrocatalytic properties of immobilized biocatalysts. In this regard, achieving direct electronic communication between the electrode and redox sites of biocatalysts eliminates the need for additional electron transfer mediators for biocatalytic applications in fuel cells and other electrochemical energy devices. In order to increase electrocatalytic currents and power in fuel cells and metal-air batteries, conductive carbon-nanostructure-modified large surface area electrodes are quite useful. Among various electrode materials, freestanding buckypapers made from carbon nanotubes have gained significance as they do not require a solid support material and thus facilitate miniaturization. In this article, we present the effect of buckypaper (BP) thickness on the electrocatalytic properties of a bilirubin oxidase (BOD) enzyme. In this study, we prepared BPs of varying thicknesses ranging from 87 µm, the minimum thickness for suitable handling with a good stability in aqueous experiments, to 380 µm. BOD was adsorbed overnight onto the BPs, mostly via hydrophobic and π-π interactions since the nanotubes used were not chemically functionalized. Furthermore, intercalation of the BOD molecules onto the nanotubes' multicylindrical network is feasible. We determined that the lower range BP thickness (<220 µm) exhibited better sigmoidal shaped electrocatalytic currents than the higher BP-thickness-based BOD biofilms with larger capacitive currents. An oxygen reduction current density of up to 3 mA cm-2 is achieved without the use of any redox mediators or tedious electrode modifications. Using the 87 µm thick BP as the representative case, we were able to obtain distinguishable peaks for all Cu sites of BOD and assign their types, T1, T2, and T3, based on the peak-width at half-maximum in anaerobic cyclic voltammograms. Our peak assignment is further supported by the appearance of dual electrocatalytic oxygen reduction waves at a higher scan rate region (>10 mV s-1) in oxygen-saturated buffer, which is identified to be driven by an â¼3.5 times faster electron transfer rate from the buckypaper to the T2/T3 center than the T1 Cu site. Findings from this study are significant for designing enzyme electrocatalytic systems and biosensors in general and fuel cells and aerobic energy storage devices in particular, where the cathodic oxygen reduction current is often inadequate.
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We present here the correlation of picomolar affinities between surface-plasmon and electrochemical immunoassays for the binding of serum glutamic acid decarboxylase 65 autoantibody (GADA), a biomarker of type 1 diabetes (T1D), to its antigen GAD-65. Carboxylated (â¼5.0%)-graphene-modified immunoassembly on a gold surface-plasmon chip or on an electrochemical array provided significantly larger binding affinity, higher sensitivity, and lower detection limits than a self-assembled monolayer surface of mercaptopropionic acid (MPA). Estimation of the relative surface -COOH groups by covalent tagging of an electroactive aminoferrocene showed that the graphenyl surface displayed a greater number of -COOH groups (9-fold) than the MPA surface. X-ray-photoelectron-spectroscopy analysis showed more C-O and CâO functionalities on the graphene-COOH surface than on the MPA surface. The graphene-COOH coating on gold exhibited â¼5.5-fold enhancement of plasmon signals compared with a similar coating on a plain glass surface. In summary, this article provides a quantitative comparison of carboxylated graphene with a mercapto-monolayer immunoassembly. Additionally, we propose that the binding-constant value can be useful as a quality-control checkpoint for reproducible and reliable production of large-scale biosensors for clinical bioassays.
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Ácido 3-Mercaptopropiônico/química , Autoanticorpos/sangue , Técnicas Eletroquímicas , Glutamato Descarboxilase/sangue , Imunoensaio , Fragmentos de Peptídeos/sangue , Ressonância de Plasmônio de Superfície , Autoanticorpos/metabolismo , Sítios de Ligação , Técnicas Biossensoriais , Glutamato Descarboxilase/metabolismo , Humanos , Fragmentos de Peptídeos/metabolismo , Propriedades de SuperfícieRESUMO
The objective of this article is to demonstrate the electrode geometric area-based scalability of pyrenyl-carbon nanostructure modification for enzyme electrocatalysis and fuel cell power output using hydrogenase anode and bilirubin oxidase cathode as the model system.
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Fontes de Energia Bioelétrica , Carbono/química , Eletrodos , Nanoestruturas/química , Hidrogenase/química , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/químicaRESUMO
Diabetes is a complex immune disorder that requires extensive medical care beyond glycemic control. Recently, the prevalence of diabetes, particularly type 1 diabetes (T1D), has significantly increased from 5% to 10%, and this has affected the health-associated complication incidences in children and adults. The 2012 statistics by the American Diabetes Association reported that 29.1 million Americans (9.3% of the population) had diabetes, and 86 million Americans (age ≥20 years, an increase from 79 million in 2010) had prediabetes. Personalized glucometers allow diabetes management by easy monitoring of the high millimolar blood glucose levels. In contrast, non-glucose diabetes biomarkers, which have gained considerable attention for early prediction and provide insights about diabetes metabolic pathways, are difficult to measure because of their ultra-low levels in blood. Similarly, insulin pumps, sensors, and insulin monitoring systems are of considerable biomedical significance due to their ever-increasing need for managing diabetic, prediabetic, and pancreatic disorders. Our laboratory focuses on developing electrochemical immunosensors and surface plasmon microarrays for minimally invasive insulin measurements in clinical sample matrices. By utilizing antibodies or aptamers as the insulin-selective biorecognition elements in combination with nanomaterials, we demonstrated a series of selective and clinically sensitive electrochemical and surface plasmon immunoassays. This review provides an overview of different electrochemical and surface plasmon immunoassays for insulin. Considering the paramount importance of diabetes diagnosis, treatment, and management and insulin pumps and monitoring devices with focus on both T1D (insulin-deficient condition) and type 2 diabetes (insulin-resistant condition), this review on insulin bioassays is timely and significant.
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Técnicas Eletroquímicas , Imunoensaio , Insulina/análise , Ressonância de Plasmônio de Superfície , Automonitorização da Glicemia , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 2/diagnóstico , HumanosRESUMO
Circulating serum nucleotide biomarkers are useful indicators for early diagnosis of cancer, respiratory illnesses, and other deadly diseases. In this work, we compared detection performances of a quartz crystal microbalance (QCM), which is a mass sensor, with that of a surface plasmon resonance (SPR) microarray for an oligonucleotide mimic of a microRNA-21 biomarker. A surface immobilized capture oligonucleotide probe was used to hybridize with the target oligonucleotide (i.e., the microRNA-21 mimic) to facilitate selective detection. To obtain ultra-low femtomolar (fM) detection sensitivity, gold nanoparticles (50 nm) were conjugated with the target oligonucleotide. We achieved detection limits of 28and 47 fM for the target oligonucleotide by the QCM and SPRi microarray, respectively. We also conducted sample recovery studies and performed matrix effect analysis. Although the QCM had a lower detection limit, the microarray approach offered better throughput for analysis of up to 16 samples. We confirmed that the designed assay was selective for the target oligonucleotide and did not show signals for the control oligonucleotide with five mismatch sites relative to the target sequence. Combination of the QCM and microarray methods that utilize the same assay chemistry on gold are useful for overcoming clinical sample matrix effects and achieving ultra-low detection of small nucleotide biomarkers with quantitative insights.
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New microarray chip strategies that are sensitive and selective and that can measure low levels of important biomarkers directly in a blood sample are significant for improving human health by allowing timely diagnosis of an abnormal condition. Herein, we designed an antibody-aptamer immunoarray chip to demonstrate simultaneous measurement of blood insulin and glycated hemoglobin (HbA1c) levels relevant to diabetic and prediabetic disorders using a surface plasmon microarray with validation by fluorescence imaging. To accomplish both surface plasmon and fluorescence imaging on the same sample, we decorated magnetite nanoparticles with quantum dots for covalent immobilization of aptamers for subsequent capture and isolation of the aptamers specific for insulin and HbA1c markers from 20-times diluted whole blood samples. Direct clinically relevant analysis, along with fluorescent imaging of the two markers, was achieved by this new immunoarray platform. The limit of detection was 4 pM for insulin and 1% for HbA1c. Examination of cross-talk using thrombin and platelet-derived growth factor confirmed that the designed immunoarray was highly selective for insulin and HbA1c. Surface plasmon kinetic analysis provided apparent binding constants of 0.24 (±0.08) nM and 37 (±3) µM, respectively, for the binding of insulin and HbA1c onto their surface immobilized monoclonal antibodies. Thus, quantitative imaging of ultralow levels of blood biomarker levels with binding kinetics is uniquely obtained in the designed immunoarray chip. In conclusion, this report demonstrates considerable significance of the developed magnetite-quantum dot-bioconjugate strategy for clinical diagnostics of whole blood biomarkers with characterization of molecular binding interactions.
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Measurement of ultra-low (e.g., parts-per-billion) levels of small-molecule markers in body fluids (e.g., serum, urine, saliva) involves a considerable challenge in view of designing assay strategies with sensitivity and selectivity. Herein we report for the first time an amperometric nano-bioelectrode design that uniquely combines 1-pyrenebutyric acid units pi-pi stacked with carboxylated multiwalled carbon nanotubes on the surface of gold screen printed electrodes for covalent attachment of NAD+ dependent formaldehyde dehydrogenase (FDH). The designed enzyme bioelectrode offered 6 ppb formaldehyde detection in 10-times diluted urine with a wide dynamic range of 10 ppb to 10 ppm. Fourier transform infrared, Raman, and electrochemical impedance spectroscopic characterizations confirmed the successful design of the FDH bioelectrode. Flow injection analysis provided lower detection limit and greater affinity for formaldehyde (apparent KM 9.6 ± 1.2 ppm) when compared with stirred solution method (apparent KM 19.9 ± 4.6 ppm). Selectivity assays revealed that the bioelectrode was selective toward formaldehyde with a moderate cross-reactivity for acetaldehyde (â¼25%) and negligible cross-reactivity toward propanaldehyde, acetone, methanol, and ethanol. Formaldehyde is an indoor pollutant, and studies have indicated neurotoxic characteristics and systemic toxic effects of this compound upon chronic and high doses of exposure. Moreover, reported chromatography and mass spectrometry methods identified elevated urine formaldehyde levels in patients with bladder cancer, dementia, and early stages of cognitive impairments compared to healthy people. Results demonstrate that pyrenyl carbon nanostructures-based FDH bioelectrode design represents novelty and simplicity for enzyme-selective electrochemical quantitation of small 30 Da formaldehyde. Broader applicability of the presented approach for other small-molecule markers is feasible that requires only the design of appropriate marker-specific enzyme systems or receptor molecules.
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Formaldeído/urina , Nanotubos de Carbono , Acetaldeído , Aldeído Oxirredutases/química , Eletrodos , Enzimas Imobilizadas/química , HumanosRESUMO
Polymer-armored enzymes loaded onto magnetic nanoparticles, as efficient nanobioreactors with enhanced properties, are described in this chapter. Polymers are useful macromolecules carrying a large number of surface charges and repeating units of desired chemical functional groups for linking enzymes onto them. Magnetic micro/nanoparticles have been widely used as enzyme carriers with the incorporation of suitable polymer layers. Synthesized iron oxide magnetic nanoparticles have been used to immobilize a peroxide-catalyzing enzyme-like heme protein: myoglobin using covalent and noncovalent strategies. The stability, scalability, and kinetics of the conjugate were studied in detail using spectroscopic and electrochemical analysis. Compared to the free myoglobin in solution, myoglobin conjugated to magnetic nanoparticles demonstrated high catalytic stability and easy recovery from the reaction medium for further use. Due to the large surface area offered by the magnetic nanoparticles, a large amount of myoglobin could be loaded with a small amount of magnetic nanoparticles. Selected examples of polymer-enzyme and polymer-magnetic nanoparticle-enzyme conjugates developed by us and others are presented in this chapter, and representative methods for making cost-effective scalable and reusable enzymatic reactors have been described.
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Enzimas Imobilizadas/química , Nanopartículas Metálicas/química , Polietilenoglicóis/química , Propilenoglicóis/química , Estabilidade Enzimática , Compostos Férricos/química , Cinética , Micelas , Nanoconjugados/química , Tamanho da Partícula , Soroalbumina Bovina/químicaRESUMO
We report single drop electroanalytical measurements of pharmaceutically and biologically relevant compounds using screen printed electrodes (SPEs) modified with carboxylated multiwalled carbon nanotubes (MWCNT-COOH) as the sensor surface. Acetaminophen, nicotine, ascorbic acid, and nicotinamide adenine dinucleotide reduced form (NADH) were detected in a single drop of solution. We show that combined polar and nonpolar interactions of analytes with -COOH functional groups and large surface area of MWCNT, respectively, allow highly sensitive analyte detection with wide dynamic range. Smaller analytes can bind to a significantly greater number of sensor sites than the bulkier analytes and offer better detection sensitivity. Results suggest that sensitivity is controlled by predominant nonpolar interactions that an analyte can undergo with the MWCNT-COOH SPE sensor surface, whereas limit of detection is controlled by the extent of polar interactions between an analyte and the sensor surface, facilitating interfacial charge transport and an electrochemical signal output. Furthermore, a combination of polar and nonpolar analyte interactions with the sensor surface shows a synergistic effect on sensitivity and detection limit. This could be a likely reason for why sensitivity does not need to always correlate with lower detection limits as variations in the interfacial interactions are critical. Application of the designed single drop method to real samples was validated by estimating the amounts of acetaminophen, nicotine, ascorbic acid, and NADH in commercially available pharmaceuticals with excellent recovery.
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We report here for the first time with quantitative details that the combination of pi-pi stacking of pyrenecarboxylic acid with chemically carboxylated multiwalled carbon nanotubes (MWNT-COOH) offers superior sensitivity compared to MWNT-COOH alone for serum insulin measurements and that this combination is broadly applicable for biosensors, drug delivery, and catalytic systems.
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Técnicas Biossensoriais/métodos , Ácidos Carboxílicos/química , Diabetes Mellitus Tipo 2/sangue , Insulina/sangue , Nanotubos de Carbono/química , Diabetes Mellitus Tipo 2/diagnóstico , HumanosRESUMO
Due to rapidly rising rates of diabetes and prediabetic conditions worldwide and the associated lethal complications, it is imperative to devise new diagnostic tools that reliably and directly measure insulin levels in clinical samples. Herein, we report a simple and sensitive direct imaging of insulin levels in diabetic patient samples using a surface plasmon resonance microarray imager (SPRi). To enhance sensitivity, we utilized magnetic nanoparticles (MNPs) to capture insulin from serum samples either directly or via a capture antibody immobilized on MNPs. The insulin-captured nanoparticles were allowed to bind surface insulin-antibody for detection from pixel intensity increase using a charge coupled device (CCD) built-in with the SPRi. We have compared the analytical figures-of-merit of the SPRi immunoarray on detecting insulin prepared in various percentages of serum solutions. A four parameter logistic model was used to obtain the best fit of microarray responses with insulin concentration and indicated the cooperative binding of insulin-nanoparticle conjugates to surface antibody in both the buffer insulin and the serum insulin conjugates with MNPs. The cooperativity effect is attributed to the greater association of magnetic nanoparticle-bound insulin molecules with increasing concentration of insulin binding to surface antibody. This is the first report of an SPRi immunoarray to accomplish clinical diagnosis of diabetic and prediabetic conditions based on insulin levels with serum matrix effect analysis and comparison between direct and sandwich insulin assay formats.
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This report investigates for the first time stability, scalability, and reusability characteristics of a protein nano-bioreactor useful for green synthesis of fine chemicals in aqueous medium extracting maximum enzyme efficiency. Enzyme catalysts conjugated with magnetic nanomaterials allow easy product isolation after a reaction involving simple application of a magnetic field. In this study, we examined a biocatalytic system made of peroxidase-like myoglobin (Mb), as a model protein, to covalently conjugate with poly(acrylic acid) functionalized magnetic nanoparticles (MNPs, 100 nm hydrodynamic diameter) to examine the catalytic stability, scalability, and reusability features of this bioconjugate. Application of the conjugate was effective for electrochemical reduction of organic and inorganic peroxides, and for both peroxide-mediated and electrocatalytic oxidation of the protein substrate 2, 2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) with greater turnover rates and product yields than Mb prepared in solution or MNP alone. Mb-attached MNPs displayed extensive catalytic stability even after 4 months of storage compared to Mb present in solution. Five- and ten-fold scale up of MNPs in the bioconjugates resulted in two- and four-fold increases in protein-catalyzed oxidation products, respectively. Nearly 40% of the initial product was present even after four reuses, which is advantageous for synthesizing sufficient products with a minimal investment of precious enzymes. Thus, the results obtained in this study are highly significant in guiding cost-effective development and efficient multiple uses of enzyme catalysts for biocatalytic, electrocatalytic, and biosensing applications via magnetic nanomaterials conjugation.
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A rapid optical microarray imaging approach for anticancer drug screening at specific cancer protein-protein interface targets with binding kinetics and validation by a mass sensor is reported for the first time. Surface plasmon resonance imager (SPRi) demonstrated a 3.5-fold greater specificity for interactions between murine double minute 2 protein (MDM2) and wild-type p53 over a nonspecific p53 mutant in a real-time microfluidic analysis. Significant percentage reflectivity changes (Δ%R) in the SPRi signals and molecular-level mass changes were detected for both the MDM2-p53 interaction and its inhibition by a small-molecule Nutlin-3 drug analogue known for its anticancer property. We additionally demonstrate that synthetic, inexpensive binding domains of interacting cancer proteins are sufficient to screen anticancer drugs by an array-based SPRi technique with excellent specificity and sensitivity. This imaging array, combined with a mass sensor, can be used to study quantitatively any protein-protein interaction and screen for small molecules with binding and potency evaluations.
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Proteínas de Neoplasias/metabolismo , Análise Serial de Tecidos , Sequência de Aminoácidos , Animais , Humanos , Proteínas de Neoplasias/química , Ligação Proteica , Teoria QuânticaRESUMO
Nanomaterial-based photoluminescence (PL) diagnostic devices offer fast and highly sensitive detection of pesticides, DNA, and toxic agents. Here we report a label-free PL genosensor for sensitive detection of Vibrio cholerae that is based on a DNA hybridization strategy utilizing nanostructured magnesium oxide (nMgO; size >30 nm) particles. The morphology and size of the synthesized nMgO were determined by transmission electron microscopic (TEM) studies. The probe DNA (pDNA) was conjugated with nMgO and characterized by X-ray photoelectron and Fourier transform infrared spectroscopic techniques. The target complementary genomic DNA (cDNA) isolated from clinical samples of V. cholerae was subjected to DNA hybridization studies using the pDNA-nMgO complex and detection of the cDNA was accomplished by measuring changes in PL intensity. The PL peak intensity measured at 700 nm (red emission) increases with the increase in cDNA concentration. A linear range of response in the developed PL genosensor was observed from 100 to 500 ng/µL with a sensitivity of 1.306 emi/ng, detection limit of 3.133 ng/µL and a regression coefficient (R(2)) of 0.987. These results show that this ultrasensitive PL genosensor has the potential for applications in the clinical diagnosis of cholera.
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Técnicas Biossensoriais , Cólera/diagnóstico , DNA Bacteriano/isolamento & purificação , Óxido de Magnésio/química , Nanoestruturas/química , Vibrio cholerae/isolamento & purificação , Cólera/microbiologia , Cólera/patologia , Sondas de DNA/síntese química , Sondas de DNA/química , DNA Bacteriano/genética , DNA Complementar/química , DNA Complementar/genética , Humanos , Limite de Detecção , Medições Luminescentes , Nanoestruturas/ultraestrutura , Hibridização de Ácido Nucleico/métodos , Processos Fotoquímicos , Espectroscopia de Infravermelho com Transformada de Fourier , Vibrio cholerae/genética , Vibrio cholerae/patogenicidadeRESUMO
HYPOTHESIS: We hypothesize that surface plasmon resonance imaging (SPRi) of interactions between small organic compounds and gold is influenced by the refractive index and chemical structures of the compounds. EXPERIMENTS: For the first time we imaged the SPR signals upon interaction of a gold surface with seven compounds representing aromatic, cyclic, short chain, and long chain carbon structures using an array format. FINDINGS: The refractive index and chemical structures of the tested compounds influenced the sensitivity of detection of the SPR microarray imager. Thus, the array methodology presented herein is suitable for studying interactions of small molecules with a gold surface.