In this reported work, we achieved high-throughput, highly sensitive fluorescent analysis using an enzyme-linked immunosorbent assay (ELISA) that employed a metallic nanoparticle (NP)-immobilized 96-well plate. The immobilization of metallic NPs on a 96-well plate effectively amplified fluorescent signals of the assay. The silver (Ag) NP-immobilized plate showed the best fluorescent enhancement effect of all plates immobilized by metal NPs. Our results demonstrate the potential of applying Ag NPs to enhance the efficiency of direct and indirect ELISA by the labeling of targets.
Extracellular vesicles (EVs) play an important role in cell-to-cell communication by carrying molecular messages that reflect physiological and pathological conditions of the parent cells. EVs have been identified in all body fluids; and among them, urine stands out as a sample that is easy and inexpensive to obtain and can be collected over time to monitor changes. Various protocols have been established to study urinary extracellular vesicles (UEVs) and they have shown great potential as a biomarker source for clinical applications, not only for urological, but also non-urological diseases. Due to the high variability and low reproducibility of pre-analytical and analytical methods for UEVs, establishing a standardized protocol remains a challenge in the field of diagnosis. Here, we review UEV studies and present the techniques that are most commonly used, those that have been applied as new developments, and those that have the most potential for future applications. The workflow procedures from the sampling step to the qualitative and quantitative analysis steps are summarized along with advantages and disadvantages of the methodologies, in order to give consideration for choosing the most promising and suitable method to analyze human UEVs.
Sensitive and specific detection of nucleic acids or proteins, which act as biomarkers, is of great importance in disease diagnosis. By combing the concept and operation of an endonuclease-assisted target-responsive amplification method and peroxidase-mimic DNAzyme generated by terminal deoxynucleotidyl transferase (TdT), a novel and facile colorimetric biosensor was developed for DNA and protein. Target DNA and thrombin were chosen as representative biomolecules. The production of cleavage fragments can only be triggered by specific target binding and the following nicking process, which do not occur spontaneously. In the signal collection part, numerous guanine-rich DNA were produced through the prolongation of cleavage fragments by TdT and formed highly effective DNAzyme with hemin. In this novel amplification method, we succeeded in realizing sensitive and specific detection of target DNA and thrombin. Under optimal conditions, target DNA can be detected as low as 1 pM, and thrombin with a detection limit of 100 pM. The method also proves the potential versatility and feasibility of TdT-generated DNAzyme in various bio-analyses.
Two novel malonyl-based chemosensors, N,N′-bis(ethyl-4′-benzoate)-1,3-propanediamide (1) and N,N′-bis(ethyl-3′-benzoate)-1,3-propanediamide (2), have been synthesized and screened towards various biologically important metal ions such as Na+, Mg2+, K+, Ca2+, Al3+, Cr3+, Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Ag+, Cd2+, Hg2+, Ti3+, and Pb2+. The emission spectral studies of both 1 and 2 displayed 84 – 91% turn-off emission responses selectively with Fe3+ ion in aqueous buffer (MeCN/H2O, 1:4, v/v, pH = 7.4) solution. Chemosensors 1 and 2 exhibited remarkable sensing ability towards Fe3+ ion over other metal ions with limit of detection (LOD) of 4.28 and 4.33 μM, respectively. The binding stoichiometry of 1 and 2 with Fe3+ ion was studied by Benesi–Hildebrand fitting, Stern–Volmer plot and Job’s plots, revealing that both chemosensors (1 – 2) bind with Fe3+ metal ion in 1:1 stoichiometric ratio with the apparent association constant (Ka) 8.90 × 103 and 11.16 × 103 M−1, respectively. Furthermore, the interactions of chemosensors (1 – 2) with metal ion were also investigated by using density functional theory (DFT) at B3LYP hybrid functional using 6-31G and LanL2DZ basis sets.
We describe here a magnetic molecular imprinted polymeric ionic liquid (MMIPIL) film by using a functionalized ionic liquid (3-vinyl-4-amino-5-imidazole carboxamide chloride, IL) and Fe3O4@Polyrutin–COOH as a functional monomer and supporting materials. The change in the direction of the charge density in the structure of MMIPIL polymer resulted in a red shift of about 100 nm for the characteristic group of –C=O. Polyrutin containing an electron-rich benzene ring and multiple hydroxyl groups not only prevented the aggregation of Fe3O4, but also benefitted to immobilize template molecules. More symmetric amino groups in the template molecules generated more hydrogen bonds and other synergistic effects between MEL and the functional monomers, which resulted in a highly-matched and highly stable MMIPIL sensor. The proposed magnetic sensor lowered the matching potential, and enhanced the signal for the detection of melamine (MEL) in milk powder. Under the optimum conditions, the MEL template molecule showed a significant linear relationship between 5.0 × 10−3 and 0.8 μg/L with a detection limit (S/N = 3) of 1.5 × 10−3 μg/L. The MMIPIL sensor showed wonderful selectivity and exhibited facile, fast and efficient results in the monitoring MEL with recoveries of between 96.5 and 108.3%.
This paper presents a study of the size distributions of colloidal nanoparticles using an online dynamic light scattering (DLS) unit with a uni-tau multi-bit correlator (UMC) combined with a centrifugal field-flow fractionation (CF3) separator. Conventionally, the FFF-UV-MALS system utilizing field-flow fractionation (FFF) combined with a UV detector and multi-angle light scattering instrument (MALS) could be used to obtain the particle size distribution of colloidal nanoparticles. Lately, DLS as a technique to measure the size distributions of colloid materials has become prevalent. However, the DLS instrument will practically measure only the large particles in a multi-modal particle mixture. Therefore, the CF3-DLS w/UMC system that was developed consisted of a CF3 unit connected to an online DLS instrument with UMC. The system could measure the volume- or number-based size distribution with highly quantitative and accurate histograms for multi-modal samples. The size distributions were validated with size distributions obtained by images of an atomic force microscope (AFM). Two types of colloidal silica nanoparticles with different distribution widths were used in this study.
Spectral pretreatments, such as background removal from Raman big data, are crucial to have a smooth link to advanced spectral analysis. Recently, we developed an automated background removal method, where we considered the shortest length of a spectrum by changing the scaling factor of the background spectrum. Here, we propose a practical way to correct the systematic error caused by noise from measurements. This correction has been realized to be more effective and accurate for automatic background removal.
Koichi HARAGUCHI, Mineshi SAKAMOTO, Akito MATSUYAMA, Megumi YAMAMOTO, Dang T. HUNG, Hiromitsu NAGASAKA, Keisuke UCHIDA, Yasunori ITO, Hitoshi KODAMATANI, Milena HORVAT, Hing M. CHAN, Matthew RAND, Ciprian M. CIRTIU, Byoung-Gwon KIM, Flemming NIELSEN, Akane YAMAKAWA, Nikolay MASHYANOV, Nikolai PANICHEV, Elena PANOVA, Tomoaki WATANABE, Naoki KANEKO, Jun YOSHINAGA, Ranny F. HERWATI, Alfrida E. SUOTH, Hirokatsu AKAGI
A certified reference material, NIMD-01, was developed for the analysis of mercury speciation in human hair. We collected the hair of Vietnamese males from a barbershop in Hanoi in 2016 and prepared 1200 bottles containing 3 g of sieved and blended hair powder. The certified value was given on a dry-mass basis, with the moisture content obtained by drying at 85°C for 4 h. Certified values with the expanded uncertainties (coverage factor, k = 2) were as follows: methylmercury, 0.634 ± 0.071 mg kg−1 as mercury; total mercury, 0.794 ± 0.050 mg kg−1; copper, 12.8 ± 1.4 mg kg−1; zinc, 234 ± 29 mg kg−1; selenium, 1.52 ± 0.29 mg kg−1. An indicative arsenic concentration of 0.17 ± 0.03 mg kg−1 was measured. Extended uncertainties were estimated by sample homogeneity, long- and short-term stabilities, and a characterization from measurements made by collaborating laboratories.
New chelating resins immobilizing carboxymethylated polyallylamine (CM-PAA) were prepared by immobilizing PAAs with some molecular weights on methacrylate resins and then carboxymethylating a part of amino groups in the PAAs using various amounts of sodium monochloroacetate. The molecular weight of PAA barely affected both the amount of PAA immobilized on the resin and the relationship between the carboxymethylation (CM) rate and the ratio of the amount of monochloroacetate used in the CM step. The selectivity of CM-PAA resin for solid-phase extraction of trace elements was almost the same as that of a resin immobilizing carboxylymethylated polyethyleneimine; 10 elements, namely Cd, Co, Cu, Fe, Mo, Ni, Pb, Ti, V, and Zn, could be quantitatively recovered over a wide pH range and alkali and alkaline earth elements were scarcely extracted under acidic and neutral conditions. The CM-PAA resin was applicable to the separation and preconcentration of the elements in a certified reference material (Waste Water, EU-L-1) and a real environmental water sample (ground water).
A fluorescent dye-labeled DNA probe was adsorbed and quenched on the monolayer of RuO2 nanosheets. Significant fluorescent recovery was observed upon the addition of complementary DNA due to desorption of the probe from the surface of the RuO2 nanosheet through duplex formation. The efficiency of fluorescence recovery was higher than that for graphene oxide, which was known as a quencher-free platform for the detection of nucleic acids in a homogeneous solution.
Yasumoto DATE, Hiroyuki MASAKI, Arata AOTA, Kazuhiro SASAKI, Yukie NAMIKI, Thomas R. GLASS, Naoya OHMURA
A simplified two-step mercury extraction procedure enabled the selective and reproducible mercury recovery from actual coal fly ash (CFA). The optimized extraction procedure involving conventional enzyme-linked immunosorbent assay (ELISA)-based immunoassay allowed the ultra-sensitive quantification of total mercury content in CFA. The total mercury content of 41 CFA samples were successfully determined using the above-mentioned method, and the results were in agreement with those obtained by standard instrumental analysis (thermal decomposition atomic absorption spectrometry) within a 15% coefficient of variation. Our method for total mercury quantification is not only simple but suitable for management of the mercury content at coal-fired electric power plants and landfill sites, which deal with large amounts of waste CFA.
Yuki TOGO, Kazunori NAKASHIMA, Wilson MWANDIRA, Satoru KAWASAKI
We developed a novel metal adsorbent composed of bio-based materials, cellulose and a protein. The approach involved the immobilization of a hexa-histidine tag (His6), which shows an affinity for an intermediate acid (metal ion) in Hard and Soft Acids and Bases (HSAB) theory, on cellulose by fusing with a carbohydrate-binding module (CBM). The results show that CBM-His6-bound cellulose has adsorption selectivity reflecting the original properties of His6. Additionally, we prepared three configurations of CBM-His6 proteins, which were subsequently immobilized on filter paper for Ni2+ ion adsorption. Of these configurations, we found that the protein containing two His6 tags at each terminus (N– and C–) of CBM exhibited the highest metal adsorption ability. Furthermore, XPS analysis confirmed the binding of Ni2+ ions on the cellulose.
This review describes recent advances in biosensors for non-invasive human healthcare applications, especially focusing on sweat analysis, along with approaches for fabricating these biosensors based on printed electronics technology. Human sweat contains various kinds of biomarkers. The relationship between a trace amount of sweat biomarkers partially partitioned from blood and diseases has been investigated by omic analysis. Recent progress in wearable or portable biosensors has enabled periodic or continuous monitoring of some sweat biomarkers while supporting the results of the omic analysis. In this review, we particularly focused on a transistor-based biosensor that is highly sensitive in quantitatively detecting the low level of sweat biomarkers. Furthermore, we showed a new approach of flexible hybrid electronics that has been applied to advanced sweat biosensors to realize fully integrated biosensing systems wirelessly connected to a networked IoT system. These technologies are based on uniquely advanced printing techniques that will facilitate mass fabrication of high-performance biosensors at low cost for future smart healthcare.
To achieve a rapid, sensitive, and economical method for the detection of ascorbic acid (AA) in the presence of Fe3+, a nitrogen and sulfur co-doped carbon dots (N,S-co-CDs) based fluorescence sensing system was developed. In this work, N,S-co-CDs were successfully synthesized via a one-step microwave-assisted method within 2.5 min using ammonium citrate and L-cysteine as precursors. The fluorescence of N,S-co-CDs was quenched (off ) by Fe3+ through a static-quenching mechanism. Subsequently, the fluorescence was recovered (on) after introducing AA into the quenched system, which was attributed to the reduction effect of AA for Fe3+. Therefore, a switch-on sensor (N,S-co-CDs/Fe3+ system) was developed for AA detection. Under optimal conditions, the limit of detection (LOD) of 2.31 μmol/L for AA was obtained over a linear range from 0 to 150 μmol/L. Furthermore, the proposed sensing method was successfully applied to detect AA in processed fruit juice with satisfactory results. The most important is that the sensor derived from a microwave-assisted method has simple and eco-friendly synthesis processes, is rapid, and has high detection efficiency. Therefore, such a switch-on sensor may be a promising candidate sensor for AA detection in processed fruit samples.
Herein, we describe the development of a novel material, “enzyme-responsive fluorescent ionic liquid”, which enabled a highly sensitive detection of alkaline phosphatase (ALP). We prepared a plasticized poly(vinyl chloride) (PVC) membrane using this new material as a plasticizer and quantified ALP in aqueous solutions. Preliminary results suggested that the PVC membrane responded to ALP at an interface between the membrane and the sample solution with anion extraction to maintain electroneutrality in the membrane phase. The developed PVC membrane showed an approximately six-times higher sensitivity than the conventional membrane, thereby demonstrating highly sensitive ALP detection. These results suggested the potential applicability of the proposed membrane for highly sensitive protein detection by using ALP-labeled antibodies.
Low aspect ratio nanopores are expected to be applied to the detection of viruses and bacteria because of their high spatial resolution. Multiphysics simulations have revealed that the ion current–time waveform obtained from low aspect ratio nanopores contains information on not only the volume of viruses and bacteria, but also the structure, surface charge, and flow dynamics. Analysis using machine learning extracts information about these analytes from the ion current–time waveform. The combination of low aspect ratio nanopores, multiphysics simulation, and machine learning has made it possible to distinguish different types of viruses and bacteria with high accuracy.
Evan L. ANDERSON, Blair K. TROUDT, Philippe BÜHLMANN
Porous glass frits are frequently used to contain the salt bridges through which reference electrodes interface samples. Prior work with widely used glass frits with 4 – 10 nm diameter pores showed that, when samples have a low electrolyte strength, electrostatic screening of sample ions by charged sites on the glass surface occurs. This creates an ion-specific phase-boundary potential at the interface between the sample and frit, and it biases the potential of the reference half-cell. Use of frits with much larger pores eliminates this problem but results in the need for frequent replenishing of the bridge electrolyte. A methodical study to determine the optimum pore size has been missing. We show here that charge screening of sample ions occurs when the pore size of nanoporous glass frits is on the order of 1 – 50 nm and samples have a low electrolyte strength. An increase in pores size to 100 nm eliminates charge screening in samples with ionic strengths in the 1.0 M to 3.3 × 10−4 M range. However, the rates of electrolyte solution flow through frits with 1, 5, 20, 50, and 100 nm pores are still low, which makes diffusion the dominant mode of ion transport into and out of these frits. Consequently, the flow of bridge electrolyte into samples is not fast enough to prevent diffusion of ions and electrically neutral components from the sample diffusing into the salt bridge, which can result in cross contamination among samples.
Local- and intermediate-range atomic structures were investigated on amorphous phases of an ordinary phase-change material, Ge2Sb2Te5 (GST), and an exotic one, Cu2GeTe3 (CGT), by using anomalous X-ray scattering close to K absorption edges of each element to find a fast amorphous-crystalline phase-change mechanism. The obtained data were analyzed by using reverse Monte Carlo modeling to obtain partial structure factors, partial pair distribution functions, and three-dimensional atomic configurations. Ring statistics were carefully examined to clarify the similarity and difference compared with the corresponding crystal structures, and it was found that amorphous GST has a number of four-membered rings indicating fragments of crystal structure, and amorphous CGT has a remarkable number of three-membered rings showing a collapse of crystal structures composed of purely six-membered rings. A persistent homology analysis was carried out and long-range ring structures of the constituent elements were observed in the amorphous phase, which may originate from fragments of crystal structures with a long-range periodicity.
Ke-Hsuan WANG, Genta WATANABE, Hayato IKEUCHI, Siyang CUI, I-Ping LIU, Kanta YAMADA, Masaaki YOSHIDA, Takeshi KAWAI
A key requirement in developing oxygen evolution reaction (OER) electrocatalysts is increasing their surface area. Herein, we report the design of a hierarchical micro/nanostructured catalyst. Based on polystyrene colloidal template electrodeposition, an ordered microcup array surrounded by nanoflakes was fabricated. The effect of the deposition time on the formation of the catalyst and the corresponding OER performance of the catalyst were investigated using scanning electron microscopy, in situ X-ray absorption fine structure (XAFS) spectroscopy, and electrochemical analysis. The in situ XAFS measurements indicate that the structure of the hierarchical structured catalyst is similar to that of γ-FeOOH. The electrochemical analysis indicates that the hierarchical catalyst has a large surface area and a low charge transfer resistance, which lead to its excellent catalytic performance for the OER. Our study provides new insights in designing high-performance OER catalysts. Moreover, the synthesized hierarchical micro/nanostructured catalyst could be used as a platform for further studies on low-cost iron-based electrocatalysts.
Soft X-ray absorption spectroscopy (XAS) involving excitation processes of a core electron to unoccupied states is an effective method to study local structures around excited C, N, and O atoms in liquid samples. Since soft X-rays are strongly absorbed by air and liquid itself, we have developed transmission-type liquid flow cells, where the absorbance of liquid samples can be easily reduced and optimized by controlling the liquid thickness. By using the transmission-mode XAS techniques, we have investigated local structures of several liquid samples such as concentration dependence of aqueous pyridine solutions and unexpected temperature-dependent structural changes in liquid benzene from the precise energy shift measurements in XAS spectra with the help of molecular dynamics simulation and inner-shell calculations. These XAS techniques are also applied to in situ/operando observation of chemical processes in solutions such as catalytic and electrochemical reactions.