RESUMEN
Atherosclerosis cardiovascular disease (ASCVD) has become one of the leading death causes in humans. Low-density lipoprotein (LDL) is an important biomarker for assessing ASCVD risk level. Thus, monitoring LDL levels can be an important means for early diagnosis of ASCVD. Herein, a novel electrochemical aptasensor for determination LDL was designed based on nitrogen-doped reduced graphene oxide-hemin-manganese oxide nanoparticles (NrGO-H-Mn3O4 NPs) integrated with clustered regularly interspaced short palindromic repeats and associated proteins (CRISPR/Cas12a) system. NrGO-H-Mn3O4 NPs not only have a large surface area and remarkable enhanced electrical conductivity but also the interconversion of different valence states of iron in hemin can provide an electrical signal. Nonspecific single-stranded DNA (ssDNA) was bound to NrGO-H-Mn3O4 NPs to form a signaling probe and was immobilized on the electrode surface. The CRISPR/Cas12a system has excellent trans-cleavage activity, which can be used to cleave ssDNA, thus detaching the NrGO-H-Mn3O4 NPs from the sensing interface and attenuating the electrical signal. Significant signal change triggered by the target was ultimately obtained, thus achieving sensitive detection of the LDL in range from 0.005 to 1000.0 nM with the detection limit of 0.005 nM. The proposed sensor exhibited good stability, selectivity, and stability and achieved reliable detection of LDL in serum samples, demonstrating its promising application prospects for the diagnostic application of LDL.
Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Sistemas CRISPR-Cas , Técnicas Electroquímicas , Grafito , Hemina , Límite de Detección , Lipoproteínas LDL , Compuestos de Manganeso , Óxidos , Compuestos de Manganeso/química , Lipoproteínas LDL/sangre , Lipoproteínas LDL/química , Humanos , Técnicas Electroquímicas/métodos , Óxidos/química , Grafito/química , Aptámeros de Nucleótidos/química , Hemina/química , Técnicas Biosensibles/métodos , ADN de Cadena Simple/química , Nanopartículas/químicaRESUMEN
Golgi protein 73 (GP73) is a new serum marker associated with early diagnosis and postoperative assessment of hepatocellular carcinoma (HCC). Herein, an electrochemical/fluorescence dual-signal biosensor was designed for determination of GP73 based on molybdenum disulfide/ferrocene/palladium nanoparticles (MoS2-Fc-PdNPs) and nitrogen-doped graphene quantum dots (NGQDs). GP73 aptamer (Apt) was labeled with NGQDs to form the NGQDs-Apt fluorescence probe. MoS2-Fc-PdNPs served not only as the fluorescence quencher but also as electrochemical enhancer. The sensing platform (NGQDs-Apt/MoS2-Fc-PdNPs) was formed based on the fluorescence resonance energy transfer (FRET) mechanism. In the presence of GP73, the specific binding of NGQDs-Apt to GP73 interrupted FRET, restoring the fluorescence of NGQDs-Apt at λex/em = 348/438 nm and enhancing the oxidation current of Fc in MoS2-Fc-PdNPs at 0.04 V through differential pulse voltammetry (DPV). Under the optimal conditions, the DPV current change and fluorescence recovery have a good linear relationship with GP73 concentration from 1.00 to 10.0 ng/mL. The calibration equation for the fluorescence mode was Y1 = (0.0213 ± 0.00127)X + (0.0641 ± 0.00448) and LOD was 0.812 ng/mL (S/N = 3). The calibration equation of the electrochemical mode was Y2 = (3.41 ± 0.111)X + (1.62 ± 0.731), and LOD of 0.0425 ng/mL (S/N = 3). The RSDs of fluorescence mode and electrochemical mode after serum detection were 1.62 to 5.21% and 0.180 to 6.62%, respectively. By combining the electrochemical and fluorescence assay, more comprehensive and valuable information for GP73 was provided. Such dual-mode detection platform shows excellent reproducibility, stability, and selectivity and has great application potential.
Asunto(s)
Carcinoma Hepatocelular , Disulfuros , Grafito , Neoplasias Hepáticas , Nanopartículas del Metal , Puntos Cuánticos , Humanos , Molibdeno , Paladio , Nitrógeno , Reproducibilidad de los Resultados , MetalocenosRESUMEN
A new sandwich-type electrochemical biosensing platform was developed by gold @polyphthalenediamine nanohybrids (AuNP@PoPD) as the sensing platform and phosphorus doped reduced graphene oxide-hemin-palladium nanoparticles (PrGO-Hemin-PdNP) as the signal amplifier for phosphatidylinositol proteoglycan 3 (GPC3). AuNP@PoPD, co-electrodeposited into the screen printed electrode with high conductivity and stability, is dedicated to assembling the primary GPC3 aptamer (GPC3Apt). The second GPC3Apt immobilized on the high conductivity and large surface area of PrGO-Hemin-PdNP was utilized as an electrochemical signal reporter by hemin oxidation (PrGO-Hemin-PdNP-GPC3Apt). In the range 0.001-10.0 ng/mL, the hemin oxidation current signal of the electrochemical aptasensor increased log-linearly with the concentration of GPC3, the lowest detection limit was 0.13 pg/mL, and the sensitivity was 2.073 µA/µM/cm2. The aptasensor exhibited good sensing performance in a human serum sample with the relative error of 4.31-8.07%. The sandwich sensor showed good selectivity and stability for detection GPC3 in human serum samples, providing a new efficient and sensitive method for detecting HCC markers.
Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Técnicas Electroquímicas , Glipicanos , Oro , Grafito , Hemina , Límite de Detección , Nanopartículas del Metal , Paladio , Glipicanos/sangre , Humanos , Técnicas Electroquímicas/métodos , Técnicas Electroquímicas/instrumentación , Aptámeros de Nucleótidos/química , Hemina/química , Grafito/química , Paladio/química , Oro/química , Técnicas Biosensibles/métodos , Nanopartículas del Metal/química , ElectrodosRESUMEN
1,5-anhydroglucitol (1,5-AG) is of considerable clinical relevance as a biochemical marker of glucose metabolism in the assessment and monitoring of diabetes. Herein, a simple colorimetric biosensor was constructed for the identification and detection of 1,5-AG by using pyranose oxidase (PROD) enzyme cascaded with reduced graphene oxide/persimmon tannin/Pt@Pd (RGO-PT/Pt@Pd NPs) nanozyme. The as-prepared RGO-PT/Pt@Pd NPs had excellent peroxidase-like activity and can be applied as a nanozyme. First, PROD enzyme reacts with the target 1,5-AG, decomposing 1,5-AG into 1,5-anhydrofuctose (1,5-AF) and H2O2. At this point, the highly catalytic RGO-PT/Pt@Pd NPs nanozyme produces a cascade with PROD enzyme which catalyzes the decomposition of H2O2 to produce O2. This in turn oxidizes the substrate 3,3',5,5'-tetramethylbenzidine (TMB) and produces a color change in the solution. Finally, the detection of 1,5-AG was achieved by measuring the absorption peak at 652 nm with an ultraviolet visible (UV-vis) spectrophotometer. Under optimal conditions, the linear operating range of the 1,5-AG enzyme cascade colorimetric sensor was 1.0-100.0 µg/mL, and the limit of detection (LOD) was 0.81 µg/mL. The proposed colorimetric biosensor was successfully applied to detect 1,5-AG in spiked human serum samples with the recoveries of 97.2-103.9% and RSDs of 1.94-4.48%. It provides a promising developmental assay for clinical detection of 1,5-AG.
Asunto(s)
Diospyros , Peróxido de Hidrógeno , Humanos , Peróxido de Hidrógeno/química , Diospyros/metabolismo , Colorimetría , Taninos , Citocromo P-450 CYP2B1 , Peroxidasa/químicaRESUMEN
Glypican-3 (GPC3), as an emerging biomarker, has been shown to be beneficial for the early diagnosis and treatment of hepatocellular carcinoma (HCC). In this study, an ultrasensitive electrochemical biosensor for GPC3 detection has been constructed based on the hemin-reduced graphene oxide-palladium nanoparticles (H-rGO-Pd NPs) nanozyme-enhanced silver deposition signal amplification strategy. When GPC3 specifically interacted with GPC3 antibody (GPC3Ab) and GPC3 aptamer (GPC3Apt), an "H-rGO-Pd NPs-GPC3Apt/GPC3/GPC3Ab" sandwich complex was formed with peroxidase-like properties which enhanced H2O2 to reduce the silver (Ag) ions in solution to metallic Ag, resulting in the deposition of silver nanoparticles (Ag NPs) on the surface of the biosensor. The amount of deposited Ag, which was derived from the amount of GPC3, was quantified by the differential pulse voltammetry (DPV) method. Under ideal circumstances, the response value was linearly correlated with GPC3 concentration at 10.0-100.0 µg/mL with R2 of 0.9715. When the GPC3 concentration was in the range from 0.01 to 10.0 µg/mL, the response value was logarithmically linear with the GPC3 concentration with R2 of 0.9941. The limit of detection was 3.30 ng/mL at a signal-to-noise ratio of three and the sensitivity was 1.535 µAµM-1cm-2. Furthermore, the electrochemical biosensor detected the GPC3 level in actual serum samples with good recoveries (103.78-106.52%) and satisfactory relative standard deviations (RSDs) (1.89-8.81%), which confirmed the applicability of the sensor in practical applications. This study provides a new analytical method for measuring the level of GPC3 in the early diagnosis of HCC.
Asunto(s)
Técnicas Biosensibles , Glipicanos , Grafito , Nanopartículas del Metal , Humanos , Técnicas Biosensibles/métodos , Carcinoma Hepatocelular , Técnicas Electroquímicas/métodos , Grafito/química , Hemina/química , Peróxido de Hidrógeno , Neoplasias Hepáticas , Nanopartículas del Metal/química , Paladio , Plata/químicaRESUMEN
Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related deaths in China. Glypican-3 (GPC3) is a specific antigen related to HCC, which is widely used in clinical detection as a reliable marker of HCC. In this paper, a highly sensitive homogeneous apatasensor was designed for GPC3 detection based on fluorescence resonance energy transfer (FRET) where the GPC3 aptamer labelled gold carbon dots (AuCDs-GPC3Apt) are used as a donor and magnetic graphene oxide (Fe3O4/GO) nanosheets are used as an acceptor. A one-step hydrothermal method was used to synthesize AuCDs to provide sufficient fluorescence. The FRET phenomenon exists between AuCDs-GPC3Apt and Fe3O4/GO, which weakens the fluorescence intensity of the whole system. When the target GPC3 is added to the FRET system, the fluorescent AuCDs-GPC3Apt binds to the GPC3 and forms a folded structure, which leads to AuCDs-GPC3Apt separation from Fe3O4/GO nanosheets. The Fe3O4/GO is then magnetically separated so that the fluorescence of free labelled AuCDs-GPC3Apt is restored. Under the optimum conditions, the fluorescence recovery rate is linearly correlated with the concentration of GPC3 (5-100 ng·mL-1) and the detection limit is 3.01 ng·mL-1 (S/N = 3). This strategy shows recoveries from 98.76 to 101.29% in real human serum samples and provides an immediate and effective detection method for the quantification of GPC3 with great potential applications for early diagnosis of HCC. A sensitive homogeneous FRET-based apatasensor was designed for GPC3 detection where the AuCDs-GPC3Apt is a donor and Fe3O4/GO nanosheets are an acceptor. The GPC3 fluorescent aptasensor combines wider output range with low cost, high specificity, and good anti-interference.
Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Carcinoma Hepatocelular , Grafito , Neoplasias Hepáticas , Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Carbono/química , Carcinoma Hepatocelular/diagnóstico , Detección Precoz del Cáncer , Transferencia Resonante de Energía de Fluorescencia/métodos , Glipicanos , Oro/química , Grafito/química , Humanos , Límite de Detección , Neoplasias Hepáticas/diagnósticoRESUMEN
A Golgi protein 73 (GP73) colorimetric biosensor based on the reduced graphene oxide-carboxymethyl chitosan-hemin/platinum@palladium nanoparticles (RGO-CMCS-Hemin/Pt@Pd NPs) with peroxidase-like activity was constructed. The RGO-CMCS-Hemin/Pt@Pd NPs with high peroxidase-like activity were successfully synthesized under mild conditions. Then, the aminylated GP73 aptamer (Apt) was bound to the RGO-CMCS-Hemin/Pt@Pd NPs to form the recognition probe. Another unmodified GP73 aptamer (AptI) was served as the capture probe. In the presence of target GP73, the capture probe and the recognition probe specifically bind to GP73 and form a RGO-CMCS-Hemin/Pt@Pd NP-Apt/GP73/AptI sandwich-type structure, which can oxidase the colorless 3,3',5,5'-tetramethylbenzidine (TMB) into blue oxTMB in the presence of H2O2. GP73 detection was achieved by measuring the peak UV absorption at 652 nm. Under the optimum conditions, the GP73 concentration was linearly related to the absorbance intensity in the range 10.0-110.0 ng/mL, and the limit of detection (LOD) was 4.7 ng/mL. The proposed colorimetric biosensor was successfully applied to detect GP73 in spiked human serum samples with recoveries of 98.2-107.0% and RSDs of 1.90-5.44%, demonstrating the excellent potential for highly sensitive GP73 detection in clinical detection. A colorimetric biosensor for visual determination of GP73 based on RGO-CMCS-Hemin/Pt@Pd NPs nanozyme with peroxidase-like activity was designed. The GP73 biosensor responses linearly from 10.0-110.0 ng/mL with LOD of 4.7 ng/mL, and shows acceptable specificity and good recovery.
Asunto(s)
Técnicas Biosensibles , Quitosano , Nanopartículas del Metal , Quitosano/química , Colorimetría , Dimaprit/análogos & derivados , Grafito , Hemina , Humanos , Peróxido de Hidrógeno/química , Nanopartículas del Metal/química , Paladio/química , Peroxidasa/química , Peroxidasas , Platino (Metal)/químicaRESUMEN
A Glypican-3 (GPC3) electrochemical aptamer nanobiosensor based on hemin/graphene nanohybrids (HGNs) peroxidase-like catalytic silver deposition and GPC3 aptamer has been constructed for the determination of GPC3. The HGNs were prepared by an one-step reduction method. Fourier transform infrared spectroscopy (FT-IR), ultraviolet spectroscopy (UV-vis), and transmission electron microscopy (TEM) were used to study the structure and morphological characteristics of the HGNs. The GPC3 electrochemical aptamer nanobiosensor was constructed using HGNs-aptamer (HGNs-Apt) as the signal probe and GPC3 aptamer as the capture probe. With the help of the catalytic action of peroxidase-like properties of HGNs, H2O2 reduces the silver (Ag) ions in solution to metallic Ag, which deposit on the surface of the electrode. The amount of deposited Ag, which was derived from the amount of GPC3, was quantified by differential pulse voltammetry (DPV). Under optimal conditions, the current response of Ag had a good positive correlation with the GPC3 concentration in the range 10.0-100.0 µg mL-1 with a correlation coefficient of 0.9958. The detection limit was 3.16 µg mL-1 at a signal-to-noise ratio of 3, and the sensitivity was calculated to be 0.807 µA µM-1 cm-2. The method is validated by analyzing spiked human serum samples with good recovery ranging from 101 to 122%. In addition, the GPC3 electrochemical aptamer nanobiosensor has acceptable selectivity, stability, and reproducibility. Graphical abstract A Glypican-3 electrochemical aptamer nanobiosensor based on hemin/graphene nanohybrids (HGNs) peroxidase-like catalytic silver deposition and GPC3 aptamer has been constructed for the determination of GPC3. The electrochemical aptamer nanobiosensor exhibits high selectivity, acceptance reproducibility, and good recovery performances.
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Técnicas Electroquímicas/métodos , Glipicanos/sangre , Nanopartículas del Metal/química , Plata/química , Catálisis , Grafito/química , Hemina/química , Humanos , Peróxido de Hidrógeno/química , Límite de Detección , Oxidación-Reducción , Reproducibilidad de los ResultadosRESUMEN
Alpha-fetoprotein (AFP) is a reliable clinical marker of hepatocellular carcinoma (HCC). A highly sensitive fluorometric aptamer nanoprobe is described for AFP detection. It is based on fluorescence resonance energy transfer (FRET) between AFP aptamer labelled with 5-carboxyfluorescein (FAM) and palladium nanoparticles (PdNPs). The PdNPs quench the green fluorescence of the FAM-AFP aptamer via interactions between nitrogen functional groups of the AFP aptamer and PdNPs. When AFP was introduced into the FAM-AFP aptamer-PdNPs FRET system, the AFP aptamer preferentially combines with AFP. This results in a conformational change and weakens the interaction between the aptamer and the PdNPs. Thus, the fluorescence of FAM recovers. The fluorescence recovery of FAM increases linearly in the 5.0-150 ng·mL-1 AFP concentration range and has a 1.4 ng·mL-1 detection limit. The assay was applied to the analysis of spiked diluted human serum. The recovery values ranged from 98.3 to 112.9%, with relative standard deviations of <1.1%. This biosensing strategy provides a reliable and ultrasensitive protocol for the quantification of biomarkers with relevant antigens and aptamers. Graphical abstract Schematic presentation of a fluorometric aptamer nanoprobe for AFP assay based on fluorescence resonance energy transfer (FRET) between AFP aptamer labelled with 5-carboxyfluorescein (FAM) and palladium nanoparticles (PdNPs).
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Fluoresceínas/química , Nanopartículas del Metal/química , Nanoestructuras/química , Paladio/química , alfa-Fetoproteínas/análisis , Animales , Aptámeros de Nucleótidos/genética , Secuencia de Bases , Estudios de Factibilidad , Transferencia Resonante de Energía de Fluorescencia , Fluorometría , Humanos , Concentración de Iones de HidrógenoRESUMEN
Sensitive and accurate detection of tumor markers is critical to early diagnosis, point-of-care and portable medical supervision. Alpha fetoprotein (AFP) is an important clinical tumor marker for hepatocellular carcinoma (HCC), and the concentration of AFP in human serum is related to the stage of HCC. In this paper, a label-free electrochemical aptasensor for AFP detection was fabricated using AFP-aptamer as the recognition molecule and thionin/reduced graphene oxide/gold nanoparticles (TH/RGO/Au NPs) as the sensor platform. With high electrocatalytic property and large specific surface area, RGO and Au NPs were employed on the screen-printed carbon electrode to load TH molecules. The TH not only acted as a bridging molecule to effectively capture and immobilize AFP-aptamer, but as the electron transfer mediator to provide the electrochemical signal. The AFP detection was based on the monitoring of the electrochemical current response change of TH by the differential pulse voltammetry. Under optimal conditions, the electrochemical responses were proportional to the AFP concentration in the range of 0.1-100.0⯵g/mL. The limit of detection was 0.050⯵g/mL at a signal-to-noise ratio of 3. The proposed method may provide a promising application of aptamer with the properties of facile procedure, low cost, high selectivity in clinic.
Asunto(s)
Aptámeros de Nucleótidos/química , Oro/química , Grafito/química , Nanopartículas del Metal/química , Tioninas/química , alfa-Fetoproteínas/análisis , Humanos , Sensibilidad y EspecificidadRESUMEN
In this study, a Fe3O4@Au-based pseudo-homogeneous electrochemical immunosensor was prepared for detection of alpha fetoprotein (AFP), a well-known hepatocellular carcinoma biomarker. The primary antibody (Ab1) was immobilized on Fe3O4@Au NPs as the capture probe. Horseradish peroxidase (HRP) and secondary antibody (Ab2) were conjugated on gold nanoparticles (GNPs) through electrostatic adsorption to form signal-amplifying labels. In the presence of AFP, a sandwich immunocomplex was formed via specific recognition of antigen-antibody in a Fe3O4@Au-basedpseudo-homogeneousreaction system. After the immunocomplex was captured to the surface of magnetic glassy carbon electrode (MGCE), the labeling HRP catalyzed the decomposition of H2O2, resulting in a substantial current for the quantitative detection of AFP. The amperometric (i-t) method was employed to record the response signal of the immunosensor based on the catalysis of the immobilized HRP toward the reduction of H2O2 with hydroquinone (HQ) as the redox mediator. Under the optimal conditions, the amperometric current response presented a linear relationship with AFP concentration over the range of 20 ng/mL-100 ng/mLwith a correlation coefficient of 0.9940, and the detection limit was 0.64 ng/mL at signal/noise [S/N] = 3. Moreover, the electrochemical immunosensor exhibited higher anti-interference ability, acceptable reproducibility and long-term stability for AFP detection.
Asunto(s)
Anticuerpos/química , Óxido Ferrosoférrico/química , Oro/química , Peroxidasa de Rábano Silvestre/química , Sondas Moleculares/química , alfa-Fetoproteínas/análisis , Técnicas Electroquímicas , Electrodos , Peroxidasa de Rábano Silvestre/metabolismo , Inmunoensayo , Nanopartículas del Metal/químicaRESUMEN
The interaction between ferrocene derivatives, such as Fc(COOH)2(λmax=286 nm), Fc(OBt)2(λmax=305 nm), Fc(Cys)(λmax=289 nm) and heme(λmax=386 nm) were studied by UV-Vis spectroscopy, respectively. The results show that, when the concentration of heme is fixed, the absorbance of heme increases with the increase of Fc(COOH)2 and Fc(Cys) concentration, the absorbance of heme almost keep the same when Fc(OBt)2 concentration increases; when the concentration of ferrocene derivatives are fixed, the absorbance of Fc(COOH)2 and Fc(Cys) also increases with the increase of heme concentration, the absorbance of Fc(OBt)2 almost keep the same when heme concentration increase. It is demonstrated that the hydrogen bonding interactions happen between Fc(COOH)2, Fc(Cys) and heme, none of Fc(OBt)2, the formation of hydrogen bonding lead to the growth of molecular chain, the bigger molecule can absorb more energy and increase the absorbance. Meanwhile, the stability of molecule is affected by the formation of hydrogen bonding, when the reaction time increases from 0.5 h to 18 h and 48 h, the absorbance at λmax=384 nm change from 2.64 to 2.53 and 2.51 with fixed concentration of Fc(COOH)2, the absorbance at λmax=384 nm change from 1.76 to 1.72 and 1.68 with fixed concentration of heme, the absorbance at λmax=397 nm change from 2.74 to 2.63 and 2.55 with fixed concentration of Fc(Cys), and the absorbance at λmax=397 nm change from 1.82 to 1.58 and 1.49 with fixed concentration of heme, respectively.
RESUMEN
Short exposures of Bacillus spores to nutrient germinants can commit spores to germinate when germinants are removed or their binding to the spores' nutrient germinant receptors (GRs) is inhibited. Bacillus subtilis spores were exposed to germinants for various periods, followed by germinant removal to prevent further commitment. Release of spore dipicolinic acid (DPA) was then measured by differential interference contrast microscopy to monitor germination of multiple individual spores, and spores did not release DPA after 1 to 2 min of germinant exposure until ~7 min after germinant removal. With longer germinant exposures, percentages of committed spores with times for completion of DPA release (Trelease) greater than the time of germinant removal (Tb) increased, while the time Tlag - Tb, where Tlag represents the time when rapid DPA release began, was decreased but rapid DPA release times (ΔTrelease = Trelease - Tlag) were increased; Factors affecting average Trelease values and the percentages of committed spores were germinant exposure time, germinant concentration, sporulation conditions, and spore heat activation, as previously shown for commitment of spore populations. Surprisingly, germination of spores given a 2nd short germinant exposure 30 to 45 min after a 1st exposure of the same duration was significantly higher than after the 1st exposure, but the number of spores that germinated in the 2nd germinant exposure decreased as the interval between germinant exposures increased up to 12 h. The latter results indicate that spores have some memory, albeit transient, of their previous exposure to nutrient germinants.
Asunto(s)
Bacillus subtilis/fisiología , Alanina/farmacología , Asparagina/farmacología , Bacillus subtilis/efectos de los fármacos , Ácidos Picolínicos/metabolismo , Esporas Bacterianas/efectos de los fármacos , Esporas Bacterianas/metabolismo , Esporas Bacterianas/fisiología , Valina/farmacologíaRESUMEN
High-precision measurements of size changes of individual bacterial spores based on ellipse fitting to bright-field images recorded with a digital camera were employed to monitor the germination of Bacillus spores with a precision of ~5 nm. To characterize the germination of individual spores, we recorded bright-field and phase-contrast images and found that the timing of changes in their normalized intensities coincided, so the bright-field images can be used to characterize spore size and refractility changes during germination. The major conclusions from this work were as follows. (i) The sizes of germinating B. cereus spores were nearly unchanged until Trelease, the time of the completion of CaDPA (a 1:1 chelate of Ca(2+) and dipicolinic acid [DPA]) release after addition of nutrient germinants. (ii) The minor axis of germinating B. cereus spores rapidly increased by ~50 nm in a few seconds right after Trelease, while the major axis was slightly decreased or unchanged. Both the minor and major axes remained unchanged for a further 30 to 45 s and then increased by 100 to 200 nm by Tlys, the time of completion of cortex lysis. (iii) Individual spores in a population showed significant heterogeneity in the timing of germination events, such as Trelease and Tlys, but also variation in size changes during germination. (iv) Bacillus subtilis wild-type spores, B. subtilis spores lacking the cortex-lytic enzyme CwlJ, and wild-type Bacillus megaterium spores showed similar kinetics of size changes during nutrient germination. The size increases in germinating spores probably result from uptake of water and cortex lysis after completion of CaDPA release.
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Bacillus subtilis/crecimiento & desarrollo , Esporas Bacterianas/química , Esporas Bacterianas/citología , Bacillus subtilis/química , Bacillus subtilis/metabolismo , Calcio/metabolismo , Cinética , Ácidos Picolínicos/metabolismo , Esporas Bacterianas/metabolismoRESUMEN
Glypican-3 (GPC3) is an essential reference target for hepatocellular carcinoma detection, follow-up and prediction. Herein, a dual-signal electrochemical aptasensor based on reduced graphene oxide-cuprous oxide (RGO-Cu2O) nanozyme was developed for GPC3 detection. The RGO-Cu2O nanoenzyme displayed excellent electron transport effect, large specific surface area and outstanding peroxidase-like ability. The differential pulse voltammetry (DPV) signal of Cu2O oxidation fraction and the chronoamperometry (i-t) signal of H2O2 decomposition catalyzed by RGO-Cu2O nanozyme were used as dual-signal detection. Under optimal conditions, the log-linear response ranges were 0.1 to 500.0 ng/mL with the limit of detection 0.064 ng/mL for DPV technique, and 0.1-50.0 ng/mL for i-t technique (detection limit of 0.0177 ng/mL). The electrochemical aptasensor has remarkably analytical performance with wide response range, low detection limit, excellent repeatability and specificity, good recovery in human serum samples. The two output signals of one sample achieve self-calibration of the results, effectively avoiding the occurrence of possible leakage and misdiagnosis of a single detection signal, suggesting that it will be a promising method in the early biomarker detection.
Asunto(s)
Técnicas Biosensibles , Cobre , Técnicas Electroquímicas , Glipicanos , Grafito , Límite de Detección , Grafito/química , Glipicanos/sangre , Glipicanos/análisis , Humanos , Técnicas Electroquímicas/métodos , Técnicas Biosensibles/métodos , Cobre/química , Aptámeros de Nucleótidos/química , Catálisis , Oxidación-Reducción , Peróxido de Hidrógeno/químicaRESUMEN
Golgi protein 73 (GP73) is a novel tumor marker in the early diagnosis and prognosis of hepatocellular carcinoma (HCC). Herein, a competitive electrochemical aptasensor for detecting GP73 was constructed using reduced graphene oxide-ferrocene-polyaniline nanocomposite (rGO-Fc-PANi) as the biosensing platform. The rGO-Fc-PANi had larger specific surface area, excellent conductivity and outstanding electroactive performance, which served as nanocarrier for GP73 aptamer (GP73Apt) binding and as redox nanoprobe for record electrical signal. Then, a complementary chain (cDNA) was fixed to the electrode by hybridization with GP73Apt. When GP73 was present, a competitive process happened among cDNA, GP73Apt and GP73, formed the GP73-GP73Apt stable chemical structure and made cDNA detach from the sensing electrode, resulting in enhancement of electrical signal. The difference in the corresponding peak current before and after the competition can be used to indicate the quantitative of GP73. Under optimal conditions, the DPV current response showed a good log-linear relationship with GP73 concentrations (0.001 â¼ 100.0 ng/mL) with a detection limit of 0.15 pg/mL (S/N = 3). It was successfully used for GP73 detection in human serum with RSDs ranging from 1.08 % to 6.96 %. Therefore, the aptasensor could provide an innovative technology platform and hold a great potential in clinical application.
RESUMEN
Breeze energy is a widely distributed renewable energy source in the natural world, but its efficient exploitation is very difficult. The conventional harvester with fixed arm length (HFA) has a relatively high start-up wind speed owing to its high and constant rotational inertia. Therefore, this paper proposes a harvester with a helix s-type vertical axis (HSVA) for achieving random energy capture in the natural breeze environment. The HSVA is constructed with two semi-circular buckets driven by the difference of the drag exerted, and the wind energy is transferred into mechanical energy. Firstly, as the wind speed changes, the HSVA harvester can match the random breeze to obtain highly efficient power. Compared with the HFA harvester, the power coefficient is significantly improved from 0.15 to 0.2 without additional equipment. Furthermore, it has more time for energy attenuation as the wind speeds dropped from strong to moderate. Moreover, the starting torque is also better than that of HFA harvester. Experiments showed that the HSVA harvester can improve power performance on the grounds of the wind speed ranging in 0.8-10.1 m/s, and that the star-up wind speed is 0.8 m/s and output peak power can reach 17.1 mW. In comparison with the HFA harvester, the HSVA harvester can obtain higher efficient power, requires lower startup speed and keeps energy longer under the same time. Additionally, as a distributed energy source, the HSVA harvester can provide a self-generating power supply to electronic sensors for monitoring the surrounding environment.
RESUMEN
1,5-Anhydroglucitol (1,5-AG) is a sensitive biomarker for real-time detection of diabetes mellitus. In this study, an electrochemical biosensor to specifically detect 1,5-AG levels based on persimmon-tannin-reduced graphene oxide-PtPd nanocomposites (PT-rGO-PtPd NCs), which were modified onto the surface of a screen-printed carbon electrode (SPCE), was designed. The PT-rGO-PtPd NCs were prepared by using PT as the film-forming material and ascorbic acid as the reducing agent. Scanning electron microscopy (SEM), transmission electron microscopy (TEM), ultraviolet-visible spectroscopy (UV-vis), and X-ray diffraction (XRD) spectroscopy analysis were used to characterise the newly synthesised materials. PT-rGO-PtPd NCs present a synergistic effect not only to increase the active surface area to bio-capture more targets, but also to exhibit electrocatalytic efficiency to catalyze the decomposition of hydrogen peroxide (H2O2). A sensitive layer is formed by pyranose oxidase (PROD) attached to the surface of PT-rGO-PtPd NC/SPCE. In the presence of 1,5-AG, PROD catalyzes the oxidization of 1,5-AG to generate 1,5-anhydrofuctose (1,5-AF) and H2O2 which can be decomposed into H2O under the synergistic catalysis of PT-rGO-PtPd NCs. The redox reaction between PT and its oxidative product (quinones, PTox) can be enhanced simultaneously by PT-rGO-PtPd NCs, and the current signal was recorded by the differential pulse voltammetry (DPV) method. Under optimal conditions, our biosensor shows a wide range (0.1-2.0 mg/mL) for 1,5-AG detection with a detection limit of 30 µg/mL (S/N = 3). Moreover, our electrochemical biosensor exhibits acceptable applicability with recoveries from 99.80 to 106.80%. In summary, our study provides an electrochemical method for the determination of 1,5-AG with simple procedures, lower costs, good reproducibility, and acceptable stability.
RESUMEN
The effective detection of biomarkers associated with hepatocellular carcinoma (HCC) is of great importance. Golgi protein 73 (GP73), a serum biomarker of HCC, has better diagnostic value than Alpha-fetoprotein (AFP) has been reported. In this paper, highly accurate fluorescence sensing platform for detecting GP73 was constructed based on fluorescence resonance energy transfer (FRET), in which nitrogen-doped graphene quantum dots (NGQDs) labelling with GP73 aptamer (GP73Apt) was used as fluorescence probe, and molybdenum disulfide @ reduced graphene oxide (MoS2@RGO) nanosheets was used as fluorescent receptors. MoS2@RGO nanosheets can quench the fluorescence of NGQDs-GP73Apt owing to FRET mechanisms. In the presence of GP73, the NGQDs-GP73Apt specifically bound with GP73 to from the deployable structures, making NGQDs-GP73Apt far away from MoS2@RGO nanosheets, blocking the FRET process, resulting in fluorescence recovery of NGQDs-GP73Apt. Under optimal conditions, the recovery intensity of fluorescence in the detection system is linearly related to the concentration of GP73 in the range of 5 ng/mL - 100 ng/mL and the limit of detection is 4.54 ng/mL (S/N = 3). Moreover, detection of GP73 was performed in human serum samples with good recovery (97.21-100.83%). This platform provides a feasible method for the early diagnosis of HCC, and can be easily extended to the detection of other biomarkers.
Asunto(s)
Aptámeros de Nucleótidos , Técnicas Biosensibles , Carcinoma Hepatocelular , Grafito , Neoplasias Hepáticas , Puntos Cuánticos , Humanos , Puntos Cuánticos/química , Grafito/química , Molibdeno/química , Nitrógeno/química , Neoplasias Hepáticas/diagnóstico , Óxidos de Nitrógeno , Aptámeros de Nucleótidos/química , Óxido Nítrico , Técnicas Biosensibles/métodosRESUMEN
OBJECTIVE: To study the value of basal serum gonadotropin levels in the diagnosis of precocious puberty (PP) in girls. METHODS: A total of 77 girls with PP were divided into central PP (CPP) (n=45) and isolated premature thelarche (IPT) groups (n=32) based on the results of gonadotropin releasing hormone (GnRH) stimulation test, which was considered the gold standard for diagnosis of PP. The two groups were compared with respect to basal serum luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels and LH/FSH ratio. The receiver operating characteristic (ROC) curve was used to analyze the accuracy of basal LH and FSH levels and LH/FSH ratio in the diagnosis of PP. RESULTS: The basal serum LH and FSH levels and LH/FSH ratio in the CPP group were significantly higher than in the IPT group (P<0.01). The basal serum LH level was positively correlated with peak LH level in the GnRH stimulation test in both groups. For diagnosis of CPP, the area under the ROC curve (AUC) for basal serum LH level was larger than for basal serum FSH level and LH/FSH ratio (P<0.05), and there was no significant difference in the AUC value between basal serum FSH level and LH/FSH ratio. When the basal serum LH level was 0.62 IU/L, there was a maximum Youden index (0.684), with 77.8% sensitivity and 90.6% specificity. When the basal serum LH level reached 1.5 IU/L, the sensitivity decreased to 31.1%, but with the highest specificity (100%). CONCLUSIONS: Basal serum LH level is superior to LH/FSH ratio and basal serum FSH level in the diagnosis of CPP, and can be used for preliminary diagnosis of PP in girls in the out-patient department, but there is some misdiagnosis and missed diagnosis. When basal serum LH level is higher than 1.5 IU/L the diagnosis of CPP can be confirmed in combination with clinical manifestation, without the need for an additional GnRH stimulation test.