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A frozen aqueous solution was integrated into a microfluidic device as a size-tunable separation field for the size-selective separation of micro/nanospheres. The width of the ice grain boundaries formed in frozen aqueous solutions could be altered by controlling the operating temperature. A freezing chamber was placed adjacent to the microfluidic channel. A sample-dispersing aqueous sucrose solution was injected into the chamber and frozen, allowing the freeze-concentrated solution (FCS) to run vertically to the microfluidic channel, where the eluting solution flows. The operating temperature can be used to control the physical interaction between the ice wall and micro/nanospheres, enabling size-selective migration. The eluted micro/nanospheres in the microchannel were passed through the eluting solution collected from the outlet. We achieved size-selective separation and collection of microspheres and nanospheres. We separated the exosomes and yeast cells to demonstrate their applicability in bioseparation. The present method is suitable not only for size-selective separation but also for evaluating the biological expression of extracellular vesicles under cryogenic conditions.
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The aggregation behavior of thermoresponsive polyelectrolytes with low charge density in aqueous solutions of organic counterions was investigated. We synthesized two thermoresponsive polyelectrolytes: anionic poly(N-isopropylacrylamide-co-(3-sulfopropyl)acrylamide potassium) (P-NIP-SPAK) and cationic poly(N-isopropylacrylamide-co-(3-acrylamidepropyl)trimethylammonium chloride) (P-NIP-AAPTAC). The polyelectrolytes remained soluble in their aqueous solutions even above the lower critical soluble temperature of P-NIP owing to the strong hydration property of the ionic groups. The aggregation occurred when organic counterions were added to the solution. In these solution systems, the concentration of counterions exceeds those of ionic groups introduced into the polyelectrolytes. The aggregation behavior is attributed to the salting-out effect of counterions accommodated near the polyelectrolyte surface by electrostatic interaction. This aggregation behavior was utilized for the charge-selective recognition of amino acids. P-NIP-SPAK aggregated only when basic amino acids were added under acidic conditions, whereas P-NIP-AAPTAC aggregated only when acidic amino acids were added under basic conditions. The results herein demonstrate that P-NIP-SPAK and P-NIP-AAPTAC have the potential to be used as charge-selective polymer sensors for amino acids without having to strictly control the experimental conditions.
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Solid-solution-type Au-Pt alloy nanoparticles (NPs) were prepared from the nanoclusters of each metal using the polymer-conjugated fusion growth method. The elemental mapping analysis showed that the mixing state of the elements in the NPs drastically changed in the narrow reaction-temperature range from 100 °C to 180 °C. For their various mixing states, the 5d-states of Au and Pt atoms in the alloy NPs were investigated on the basis of the white line intensities of X-ray absorption near edge structure (XANES). Then, the 5d-states of Au and Pt atoms in a model crystalline ordered alloy structures were investigated on the basis of the theoretically calculated XANES spectra using density functional theory (DFT) in the whole composition range. The DFT calculation showed that the changes in the absorption spectra near the Pt and Au edges are caused by the change in the occupation of the Pt 5d-states and the orbital hybridisation of the Au 5d-states with the 5d-states of neighbouring Pt atoms around an Au atom, respectively.
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A novel Raman scattering enhancement was discovered using colloid nanoparticles conjugated with an amine-based copolymer. The interaction potential surface between Raman scattering enhancing nanoparticles was clarified by combining a small-angle scattering method and a model-potential-free liquid-state theory as an in situ observation in the solution state. The potential surface indicates that the most stable position is located around 0.9 nm from the particle surface, suggesting the existence of a nanogap structure between the nanocomposites. The change in Raman scattering enhancement was also acquired during the dispersion process of the aggregated nanocomposites through a glutathione-triggered nanosensing reaction.
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Resinas Acrílicas/química , Nanocompostos/química , Análise Espectral Raman/métodos , Resinas Acrílicas/síntese química , Glutationa/química , Ouro/química , Nanopartículas Metálicas/química , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Interaction between the thermoresponsive polymer poly(N-isopropylacrylamide) (P-NIP) and sodium dodecyl sulfate (SDS) both above and below its phase transition temperature was examined under dilute conditions. Above the lower critical solution temperature (LCST) of P-NIP (32 °C), 0.01 wt % P-NIP specifically interacted with 1.0 × 10(-5) mol/L SDS to form a precipitate. However, when SDS was added at concentrations above or below 1.0 × 10(-5) mol/L, the P-NIP solution remained clear above the LCST. A fluorometric probe, N-phenyl-naphthalene, indicated that the hydrophobicity of the aggregates composed of P-NIP and SDS changed at an SDS concentration of 1.0 × 10(-5) mol/L. Although the hydrophobicity of the precipitate was similar to that of P-NIP alone at less than 1.0 × 10(-5) mol/L, it approached that of SDS homomicelles as the SDS concentration increased above 1.0 × 10(-5) mol/L. Dynamic light scattering and turbidimetry studies showed no P-NIP phase transition above an SDS concentration of 1.0 × 10(-5) mol/L, which is much lower than the reported critical association concentration (CAC) of SDS with P-NIP. This indicates that P-NIP interacted with SDS above the LSCT at much lower SDS concentration than the reported CAC.
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Therma-Max™ LSA Streptavidin is a thermoresponsive magnetic nanoparticle (TMNP). It can be introduced conveniently to molecular recognition groups by avidin-biotin interaction. In this study, we demonstrated the detection of nickel(II) ions by the magnetic separation of TMNP induced by their phase transition under microflow. The NTA-tagged TMNP solution mixed with a Ni2+ sample was introduced into a microchannel with a well structure. Moreover, the sample was heated to induce the thermally induced aggregation of TMNP. The Ni-capturing TMNP were trapped in the well by magnetic fields. The supernatant was removed from the outlet, and a dimethylglyoxime (DMG) solution was introduced into the device for colorimetric detection in the well. Because DMG has a higher stability constant with Ni2+, sensitive colorimetric detection of Ni2+ can be achieved in devices where the sample volume, e.g., optical pathlength, is short. To demonstrate the feasibility of the proposed method, a recovery test was conducted using a commercially available cosmetic sample. Therein, complete collection was achieved.
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BACKGROUND: Psychrophiles can survive under cryogenic conditions because of various biomolecules. These molecules interact with cells, ice crystals, and lipid bilayers to enhance their functionality. Previous studies typically measured these interactions by thawing frozen samples and conducting biological assays at room temperature; however, studying these interactions under cryogenic conditions is crucial. This is because these biomolecules can function at lower temperatures. Therefore, a platform for measuring chemical interactions under sub-zero temperature conditions must be established. RESULTS: The chemical interactions between biomolecules under sub-zero temperature conditions were evaluated within ice grain boundaries with a channel-like structure, which circumvents the need for thawing. An aqueous solution of sucrose was frozen within a microfluidic channel, facilitating the formation of freeze-concentrated solutions (FCSs) that functioned as size-tunable electrophoretic fields. Avidin proteins or single-stranded DNA (ssDNA) were introduced into the FCS in advance. Probe micro/nanospheres whose surfaces were modified with molecules complementary to the target analytes were introduced into the FCS. If the targets have functionalities under sub-zero temperature conditions, they interact with complementary molecules. The chemical interactions between the target molecules and nanospheres led to the aggregation of the particles. The size tunability of the diameter of the FCS channels enabled the recognition of aggregation levels, which is indicative of interaction reactivity. The avidin-biotin interaction and ssDNA hybridization served as models for chemical interactions, demonstrating interactivity under sub-zero temperature conditions. The results presented herein suggest the potential for in situ measurement of biochemical assays in the frozen state, elucidating the functionality of bio-related macromolecules at or slightly below 0 °C. SIGNIFICANCE: This is the first methodology to evaluate chemical interactions under sub-zero temperature conditions without employing the freeze-and-thaw process. This method has the advantage of revealing the chemical interactions only at low temperatures. Therefore, it can be used to screen and evaluate the functionality of cryo-related biomolecules, including cold-shock and antifreeze proteins.
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Temperatura Baixa , Eletroforese , DNA de Cadeia Simples/química , DNA de Cadeia Simples/análise , Gelo/análise , CongelamentoRESUMO
Kinetic studies on the photochemical quenching reaction of the tris(2,2'-bipyridyl) ruthenium(II) complex ([Ru(bpy)3]2+) in water-glycerol binary media were conducted based on the Einstein-Smoluchowski (E-S) theory. Dynamic and static quenching behaviors were analyzed by comparing results from time-resolved spectroscopy and emission spectroscopy. While the dynamic quenching reaction aligns well with the E-S theory, static quenching was observed, leading to a notable increase in the overall photoquenching reaction rate constant. Employing chromatography and infrared spectroscopy, we correlated the microscopic molecular structure of the binary solvent system and the solvation environment around the emitters with the reaction mechanism. This correlation was found to correspond to ion pair formation and the confinement effect of the emitter, respectively.
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Acquisition protocol of absorption spectra at nanoliter spaces from the RGB values preserved in video data at 10 ms intervals was established using the principal-component-analysis-based RGB-conversion method. Proton behavior was monitored using a camera to acquire the video footage to monitor colorimetric change in the nanoliter space. The RGB values observed in the video were converted into a score vector using a conversion matrix. A linear combination of the predetermined loading vectors with the score values was calculated to reproduce the absorption spectra. The reproduced absorption spectra correlated well with those acquired using a conventional spectrophotometer during a short period. This method was applied to monitor the proton diffusion from a single cationic ion-exchange resin into hydrogels at low concentrations. The rapid acquisition and quick response of this method may enable the monitoring of the initial diffusion of protons, which is challenging with conventional spectrophotometry and electrochemical approaches.
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A stencil printing method utilizing sticky notes, a thermal transfer ink ribbon, and office appliances for paper-based analytical device (PAD) fabrication was proposed. A sticky note was attached to a filter paper, and a mask pattern was cut using a cutting machine. A commercially available thermal ink ribbon was then placed over the mask and laminated. We have characterized the fabricated devices. This approach could be used for the fast and mass prototyping of PADs using simple office appliances with no need for a wax printer.
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Total protein concentrations in the aqueous solutions were determined from the absorption spectra reproduced from smartphone-captured digital color images. We employed two different procedures for protein determination: the pyrogallol red molybdate method and Bradford's method. The principal-component-analysis-based reproduction process, which was previously reported by our research group, enabled the conversion of RGB values to score values for a linear combination of loading vectors to generate reproduced absorption spectra. The reproduced spectra were identical to those measured using a commercially available spectrophotometer. The total protein assays of commercial soymilk and human serum samples were carried out with both coloration reagents, and the obtained results were in good agreement with those attained using a conventional spectrophotometer. These results show that the proposed method enables smartphone-based ratiometric analysis of real samples without requiring any monochromating equipment.
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Bioensaio , Smartphone , Colorimetria/métodos , Humanos , EspectrofotometriaRESUMO
The absorption spectra of bromothymol blue (BTB) solution introduced in microfluidic devices were reproduced by principal component analysis (PCA)-based xyz-spectrum conversion methods for geometric mapping of the pH values of fluids. We fabricated PDMS-made microfluidic devices with a channel depth of 1 mm to overcome the lower detection limits of transmittance image acquisition. Aqueous solutions of pH indicators under various pH conditions were hydrodynamically introduced into the channel, and RGB values of the region of interest (ROI) were obtained via image analysis. The xyz values were then converted into absorption spectral data of the pH indicator using the PCA-based spectral reproduction previously proposed by the authors. The high reproducibility of the spectra was confirmed to be comparable to that of the conventional method using a spectrophotometer. We applied the present method to elucidate the pH gradient at an aqueous biphasic interface in the microfluidic channels generated by contacting multiple laminar flows of two or three buffered solutions. We confirmed that the pH gradient ranged from approximately 70 to 140 µm, which is consistent with the results reported using other approaches. The results demonstrate the applicability of the present method to the fluctuation field in micro/nanospaces to acquire spectrophotometric information in the order of milliseconds without monochromating equipment.
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Dispositivos Lab-On-A-Chip , Microfluídica , Concentração de Íons de Hidrogênio , Análise de Componente Principal , Reprodutibilidade dos TestesRESUMO
This article describes the glutathione-triggered disassembly of gold nanocomposites composed of gold cores and water-soluble copolymers [poly(N-n-isopropylacrylamide-co-acryloyldiethyletriamine)] attached to the surfaces of gold cores. The gold nanocomposites exhibit a bluish purple color because of the assembled gold cores that are conjugated with the diethylenetriamine groups incorporated into the copolymers. Glutathione added to the gold nanocomposite solution adsorbs onto the surface of the gold cores to liberate diethylenetriamine groups, resulting in spontaneous disassembly that changes the color of the solution to a reddish shade. Increasing the glutathione concentration facilitates the spontaneous disassembly of the gold nanocomposites. For the determination of glutathione, the colorimetric change of the gold nanoparticles is quantified with the a* value of the L*a*b* color coordinates defined by the CIE (Commission Internationale de l'Eclairage) chromaticity diagram. A linear relationship between the a* value and the glutathione concentration of up to 6 x 10(-6) mol/L is obtained 15 min after the addition of glutathione that has a detection limit (defined as 3sigma) of 2.9 x 10(-8) mol/L. The colorimetric assay is successfully applied to the determination of glutathione in eye drops and health supplements.
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Glutationa/análise , Ouro/química , Nanocompostos/química , Polímeros/química , Água/química , Colorimetria , Glutationa/química , Modelos Moleculares , Conformação Molecular , Soluções Oftálmicas/química , Solubilidade , Compostos de Sulfidrila/química , TemperaturaRESUMO
Absorption spectra of pH indicators in aqueous solutions were reproduced from RGB values of microscopic images utilizing principal component analysis (PCA) and linear algebraic treatments. The reproduction of absorption spectra comprises the following three steps: (1) determining the loading spectra by PCA, (2) determining the conversion matrix from the RGB values to the score vectors, and (3) reproducing the absorption spectra by linear combination of the loading spectra and the score vectors. The reproducibility of the absorption spectra was demonstrated by employing bromothymol blue and methyl red solutions as pH indicators. The reproduced spectra of both indicators were in good agreement with the spectra measured with a conventional spectrophotometer. The pKa values of both indicators calculated from the reproduced spectra are in good agreement with those obtained from the spectrophotometric spectra and the literature values, confirming validity of the reproduction. This approach was applied to measure pH of freeze concentrated solutions in micro drains formed in ice. A change in pH was successfully observed on freezing and was compared with that reported in previous literature. Since this method does not necessitate the use of grating systems, spectral changes can be traced in milliseconds; this elucidates the phenomena occurring in fluctuating fields.
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The chemical interaction between antifreeze proteins (AFPs) and ice crystals is evaluated via electrophoresis of AFP-anchored microparticles in fluidic channels formed in frozen aqueous sucrose. Straight fluidic channels are created in a flat glass chamber connecting two Ag/AgCl electrodes. This configuration allows us to estimate an electric field strength exerted on probe particles migrating along the channel. When the channel width is comparable to the particle size, the particle is immobile because of the resistance force induced by the interaction with the ice wall. However, when the overall electrophoretic force surpasses the resistance force, the microsphere starts to migrate. From the threshold electric field strengths determined for unmodified and AFP-modified particles, the resistance forces for the chemical interaction between AFPs and ice wall are estimated.
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Proteínas Anticongelantes/química , Gelo , Sacarose/química , Sítios de Ligação , Cristalização , Eletricidade , Eletroforese , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
The present dataset is related to the research paper entitled "Reproducing Absorption Spectra of pH Indicators from RGB Values of Microscopic Images" (Inagawa et al., 2020). The dataset contains microscopic images of aqueous methyl orange (MO), absorption spectra acquired with a spectrophotometer, loading spectra and calculation sheets for reproducing absorption spectra of aqueous MO from their RGB values of the microscopic images. The microscopic transmission images of the standard MO solutions at various pH conditions were acquired with a CMOS camera equipped with an invert microscope. Meanwhile, the loading spectra were obtained by principle component analysis of a series of absorption spectra of the standard solutions. The conversion matrix from RGB values in a region of interests (ROI) to score values were linear-algebraically determined from the RGB values and score values of the standard solutions. The absorption spectra of the sample solutions of which pH conditions are unknown were then reproduced by calculating the linear combination of the loading spectra with the score values obtained from the conversion process. Herein, the absorption spectra of MO are reproduced at various pH and ROI conditions.
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We describe a simple, yet selective cysteine sensor based on gold nanoparticles (AuNPs) conjugated with thermoresponsive copolymers, the carboxyl groups of which are incorporated. Copolymer-conjugated AuNPs, used as the cysteine sensor, in a solution form sediment when cysteine is added. Heating followed by cooling the solution induces phase transitions of the thermoresponsive copolymers, resulting in an acceleration of sedimentation of the copolymer-conjugated AuNPs. The absorbance of supernatants at 520 nm, which are ascribed to a surface plasmon band of discrete AuNPs, decays with increasing concentration of cysteine. Sedimentation of the copolymer-conjugated AuNPs is specific to cysteine. The addition of other popular amino acids, or ascorbic acid, causes no sedimentation of the AuNPs. The relationship between the absorbance of the supernatant at 520 nm and the cysteine concentration provides a sigmoidal profile at a concentration range between 1 x 10(-6) to 6 x 10(-6) mol dm(-3). The determination of cysteine in a supplement is achieved using an inflection point on the sigmoidal profiles.
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Tris(2-methyl-8-quinolinolato)iron(III) was proposed as a sensitive spectrophotometric silanol-detecting probe based on the coordination ability of silanol groups on the surface of octadecylsylanized silica gel (ODS silica gel) for reversed-phase high-performance liquid chromatography. A peak of the iron(III) complex on a chromatogram abruptly collapsed as the silanol content in an ODS column increased, indicating that the iron(III) complex could sense trace amounts of silanol groups. The change of the peak parameters, such as the peak height and the peak area was highly related to the output of some nitrogen-containing compounds used as silanol-detecting probes as a function of the silanol content in an ODS column. The response of the peak height of the iron(III) complex to the silanol content was much more sensitive than the response of the nitrogen-containing probes, and was comparable to that of tris(2-methyl-8-quinolinolato)gallium(III), which had been proposed as a fluorometric silanol-detecting probe based on the coordination ability of the silanol groups.
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The peak shape of tris(2-methyl-8-quinolinolato)gallium(III) by reversed-phase high-performance liquid chromatography was found to be very sensitive to trace amounts of silanol groups on the surface of octadecylsylanized silica gel (ODS silica gel). The variation of the peak of the gallium(III) complex can be used as a probe of the residual silanol groups in an ODS column. The chromatographic peak parameters of the complex were compared with the silanol activities output by some silanol-detecting tests using nitrogen-containing compounds as probes. The comparison was performed with several commercially available ODS columns and laboratory-packed columns in which the amount of silanol groups was controlled by mixing fully endcapped ODS materials and a non-endcapped ODS material. The peak height was the most effective parameter among the peak parameters, and much more sensitive than the silanol-detecting tests using nitrogen-containing compounds, in detecting a trace amount of silanol groups that could not be detected by other silanol-detecting tests.
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Corantes Fluorescentes/química , Gálio/química , Compostos Organometálicos/química , Quinolinas/química , Silanos/análise , Cromatografia Líquida de Alta Pressão/métodos , Sensibilidade e EspecificidadeRESUMO
Cadmium species in river water were kinetically extracted with dithizone by varying the extraction time. The obtained extraction curve showed a three-stage stepwise profile that reflected the rate of the ligand substitution reaction between the dithizone and cadmium species. Corresponding to each stage, we divided these extracted cadmium species into three groups: "highly labile", "moderately labile" and "slowly labile" species.