Tomokazu Matsue

Tohoku University, Japan

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Publications (284)771.9 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: We have developed a novel method for detection of endotoxin with extra-high sensitivity by using substitutional stripping voltammetry (SSV). In this method, a p-aminophenol (pAP) conjugated peptide (Boc-Leu-Gly-Arg-pAP; LGR-pAP) was used as a substrate for a protease, which is activated at the last step of the endotoxin-induced Limulus amebocyte lysate (LAL) cascade reaction. Extra-highly sensitive detection of pAP liberated by the endotoxin-induced LAL reaction was successfully realized with SSV, based on the accumulation of an amperometric signal owing to exchange of the oxidation current of pAP generated at an electrode in a reaction cell with silver deposition on another electrode in a deposition cell. This reaction is driven by the difference in the redox potential between pAP/quinoneimine and silver/silver ion. The amount of the deposited silver is quantified by anodic stripping voltammetry (ASV). This SSV-based endotoxin assay was performed with a chip device comprising two cells, each of which was connected via a liquid junction made of Vycor® glass. The reaction cell and the deposition cell contained a standard endotoxin sample with LAL regents containing LGR-pAP and AgNO3 solution, respectively. After the cells were electrically connected for 60 min, ASV was conducted in the deposition cell to quantify the total electrical charge derived by the oxidation of free pAP in the reaction cell. The ASV signal increased with the increase of the endotoxin concentration in the sample solution in the range of 0.5-1000 EU L(-1).
    The Analyst 08/2014; · 4.23 Impact Factor
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    ABSTRACT: In this manuscript we demonstrate the rapid formation of three-dimensional (3D) embryonic stem cell (ESC) aggregates with controllable sizes and shapes in hydrogels using dielectrophoresis (DEP). The ESCs encapsulated within a methacrylated gelatin (GelMA) prepolymer were introduced into the DEP device and, upon applying an electric field and crosslinking the GelMA hydrogel, formed 3D ESC aggregates. Embryonic bodies (EBs) fabricated using this method showed high cellular viability and pluripotency. The proposed technique enables production of EBs on a large-scale and in a high-throughput manner for potential cell therapy and tissue regeneration applications.
    Lab on a Chip 07/2014; · 5.70 Impact Factor
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    ABSTRACT: This paper reports a novel approach for the simple detection of cell apoptosis using an electrochemical technique. This method uses caspase-3 activity as an indicator of apoptosis. Caspase-3 activity was detected with differential plus voltammetry (DPV) as an alternative to conventional spectrometry. In this method, p-nitroaniline (pNA) released from Asp-Glu-Val-Asp-pNA by caspase-3 enzyme reaction was measured with DPV by using a glassy carbon electrode. Using this method, we successfully detected cell apoptosis occurring inside living HepG2 cells without the need for a cell lysis step. This method provides an easy assay procedure and, more importantly, allows a live cell apoptosis detection format. This novel electrochemical apoptosis assay using living cells instead of typically used cell lysates will expand the applicable range of the apoptosis assay to include cell activity assays for drug discovery and cell transplantation medicine.
    Analytical Chemistry 05/2014; · 5.70 Impact Factor
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    ABSTRACT: In the present study, gene analysis using siRNAs for cell differentiation of embryonic stem (ES) cells was performed using an LSI-based amperometric sensor array (Bio-LSI). CITED2 and WNT11 were silenced using siRNA, and then the effect of these genes on the differentiation of ES cells into cardiomyocytes was evaluated. For the evaluation, endogenous ALP activity and sarcomeric α-actinin were electrochemically detected using the Bio-LSI system. These results show the crucial role of these genes in determining the fate of differentiation of ES cells. These results also show that the Bio-LSI is a great tool for cell analysis.
    Analytical methods 04/2014; · 1.86 Impact Factor
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    ABSTRACT: We propose a novel application of dielectrophoresis (DEP) to make three-dimensional (3D) methacrylated gelatin (GelMA) hydrogels with gradients of micro- and nanoparticles. DEP forces were able to manipulate micro- and nanoparticles of different sizes and materials (i.e., C2C12 myoblasts, polystyrene beads, gold microparticles, and carbon nanotubes) within GelMA hydrogels in a rapid and facile way and create 3D gradients of these particles in a microchamber. Immobilization of drugs, such as fluorescein isothiocyanate-dextran (FITC-dextran) and 6-hydroxydopamine (6-OHDA), on gold microparticles allowed us to investigate the high-throughput release of these drugs from GelMA-gold microparticle gradient systems. The latter gradient constructs were incubated with C2C12 myoblasts for 24h to examine the cell viability through the release of 6-OHDA. The drug was released from the microparticles in a gradient manner, inducing a cell viability gradient. This novel approach to create 3D chemical gradients within hydrogels is scalable to any arbitrary length scale. It is useful for making anisotropic biomimetic materials and high-throughput platforms to investigate cell-microenvironment interactions in a rapid, simple, cost-effective, and reproducible manner.
    Biosensors & bioelectronics 03/2014; 59C:166-173. · 5.43 Impact Factor
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    ABSTRACT: A new local redox cycling-based electrochemical (LRC-EC) device integrated with many electrochemical sensors has been developed into a small chip device. The LRC-EC chip device was successfully applied for the detection of alkaline phosphatase and horseradish peroxidase activity based on substrate generation/chip collection (SG/CC) and extended feedback modes, respectively. The new imaging approach with extended feedback mode was particularly effective for sharpening of the image, because this mode used feedback signals and minimizes the undesired influence of diffusion. The LRC-EC chip device is considered to be a useful tool for bioanalysis.
    Analytical Chemistry 03/2014; · 5.70 Impact Factor
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    ABSTRACT: A rapid, ultra-sensitive, and practical label-free impedimetric immunoassay for measuring trace levels of total polychlorinated biphenyls (PCBs) in insulating oil was developed. First, we developed a novel monoclonal antibody (RU6F9) for PCBs by using a designed immunogen and characterized its binding affinity for a commercial mixtures of PCBs, and its main congeners. A micro comb-like gold electrode was fabricated, and the antibody was covalently immobilized on the electrode through a self-assembled monolayer formed by dithiobis-N-succinimidyl propionate. The antigen-binding event on the surface of the functionalized electrode was determined as the change in charge transfer resistance by using electrochemical impedance spectroscopy. The resulting impedimetric immunoassay in aqueous solution achieved a wide determination range (0.01-10 µg/L) and a low detection limit (LOD) of 0.001 µg/L, which was 100-fold more sensitive than a conventional flow-based immunoassay for PCBs. By combining the impedimetric immunoassay with a clean-up procedure for insulating oil utilizing a multilayer clean-up column followed by DMSO partitioning, an LOD of 0.052 mg/kg-oil was achieved, which satisfied the Japanese regulation criterion of 0.5 mg/kg-oil. Finally, the immunoassay was employed to determine total PCB levels in actual used insulating oils (n = 33) sampled from a used transformer containing trace levels of PCBs, and the results agreed well with the Japanese official method (HRGC/HRMS).
    Analytical Chemistry 02/2014; · 5.70 Impact Factor
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    ABSTRACT: As a complementary tool to nanofluidics, biomolecular based transport is envisioned for nanotechnological devices. We report a new method for guiding microtubule shuttles on multi-walled carbon nanotube tracks, aligned by dielectrophoresis on a functionalized surface. In the absence of electric field and in fluid flow, alignment is maintained. The directed translocation of kinesin propelled microtubules has been investigated using fluorescence microscopy. To our knowledge, this is the first demonstration of microtubules gliding along carbon nanotubes.
    Nano Letters 01/2014; · 13.03 Impact Factor
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    ABSTRACT: NAD(P)H: quinone oxidoreductase (NQO) activity of single HeLa cells were evaluated by using the menadione-ferrocyanide double mediator system combinded with scanning electrochemical microscopy (SECM).
    Analytica Chimica Acta. 01/2014;
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    ABSTRACT: A flexible sensor based on SU-8 photoresist was fabricated and its electrochemical performance was investigated using cyclic voltammetry. The device consisted of interdigitated array (IDA) electrodes on an SU-8 layer. It exhibited a clear electrochemical response during redox cycling of ferrocenemethanol at the IDA electrodes. Since the device was flexible, it could be inserted into a narrow bent space to monitor electrochemical responses. The observed electrochemical behavior was found to be consistent with that predicted by simulations based on redox compound diffusion.
    Analytical Sciences 01/2014; 30(2):305-9. · 1.57 Impact Factor
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    ABSTRACT: Biological scaffolds with tunable electrical and mechanical properties are of great interest in many different fields, such as regenerative medicine, biorobotics, and biosensing. In this study, dielectrophoresis (DEP) was used to vertically align carbon nanotubes (CNTs) within methacrylated gelatin (GelMA) hydrogels in a robust, simple, and rapid manner. GelMA-aligned CNT hydrogels showed anisotropic electrical conductivity and superior mechanical properties compared with pristine GelMA hydrogels and GelMA hydrogels containing randomly distributed CNTs. Skeletal muscle cells grown on vertically aligned CNTs in GelMA hydrogels yielded a higher number of functional myofibers than cells that were cultured on hydrogels with randomly distributed CNTs and horizontally aligned CNTs, as confirmed by the expression of myogenic genes and proteins. In addition, the myogenic gene and protein expression increased more profoundly after applying electrical stimulation along the direction of the aligned CNTs due to the anisotropic conductivity of the hybrid GelMA-vertically aligned CNT hydrogels. We believe that platform could attract great attention in other biomedical applications, such as biosensing, bioelectronics, and creating functional biomedical devices.
    Scientific Reports 01/2014; 4:4271. · 5.08 Impact Factor
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    ABSTRACT: We propose a novel application of dielectrophoresis (DEP) to make three-dimensional (3D) methacrylated gelatin (GelMA) hydrogels with gradients of micro- and nanoparticles. DEP forces were able to manipulate micro- and nanoparticles of different sizes and materials (i.e., C2C12 myoblasts, polystyrene beads, gold microparticles, and carbon nanotubes) within GelMA hydrogels in a rapid and facile way and create 3D gradients of these particles in a microchamber. Immobilization of drugs, such as fluorescein isothiocyanate–dextran (FITC–dextran) and 6-hydroxydopamine (6-OHDA), on gold microparticles allowed us to investigate the high-throughput release of these drugs from GelMA-gold microparticle gradient systems. The latter gradient constructs were incubated with C2C12 myoblasts for 24 h to examine the cell viability through the release of 6-OHDA. The drug was released from the microparticles in a gradient manner, inducing a cell viability gradient. This novel approach to create 3D chemical gradients within hydrogels is scalable to any arbitrary length scale. It is useful for making anisotropic biomimetic materials and high-throughput platforms to investigate cell-microenvironment interactions in a rapid, simple, cost-effective, and reproducible manner.
    Biosensors and Bioelectronics. 01/2014; 59:166–173.
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    ABSTRACT: Tissue engineering (TE) is a multidisciplinary research area that combines medicine, biology, and material science. In recent decades, microtechnology and nanotechnology have also been gradually integrated into this field and have become essential components of TE research. Tissues and complex organs in the body depend on a branched blood vessel system. One of the main objectives for TE researchers is to replicate this vessel system and obtain functional vascularized structures within engineered tissues or organs. With the help of new nanotechnology and microtechnology, significant progress has been made. Achievements include the design of nanoscale-level scaffolds with new functionalities, development of integrated and rapid nanotechnology methods for biofabrication of vascular tissues, discovery of new composite materials to direct differentiation of stem and inducible pluripotent stem cells into the vascular phenotype. Although numerous challenges to replicating vascularized tissue for clinical uses remain, the combination of these new advances has yielded new tools for producing functional vascular tissues in the near future.
    Journal of Nanoscience and Nanotechnology 01/2014; 14(1):487-500. · 1.15 Impact Factor
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    ABSTRACT: Electrical properties of cells determine most of the cellular functions, particularly ones which occur in the cell's membrane. Manipulation of these electrical properties may provide a powerful electrotherapy option for the treatment of cancer as cancerous cells have been shown to be more electronegative than normal proliferating cells. Previously, we used an electrical impedance sensing system (EIS) to explore the responses of cancerous SKOV3 cells and normal HUVEC cells to low intensity (<2 V/cm) AC electric fields, determining that the optimal frequency for SKOV3 proliferation arrest was 200 kHz, without harming the non-cancerous HUVECs. In this study, to determine if these effects are cell type dependant, human breast adenocarcinoma cells (MCF7) were subjected to a range of frequencies (50 kHz-2 MHz) similar to the previously tested SKOV3. For the MCF7, an optimal frequency of 100 kHz was determined using the EIS, indicating a higher sensitivity towards the applied field. Further experiments specifically targeting the two types of cancer cells using HER2 antibody functionalized gold nanoparticles (HER2-AuNPs) were performed to determine if enhanced electric field strength can be induced via the application of nanoparticles, consequently leading to the killing of the cancerous cells without affecting non cancerous HUVECs and MCF10a providing a platform for the development of a non-invasive cancer treatment without any harmful side effects. The EIS was used to monitor the real-time consequences on cellular viability and a noticeable decrease in the growth profile of the MCF7 was observed with the application of the HER2-AuNPs and the electric fields indicating specific inhibitory effects on dividing cells in culture. To further understand the effects of the externally applied field to the cells, an Annexin V/EthD-III assay was performed to determine the cell death mechanism indicating apoptosis. The zeta potential of the SKOV3 and the MCF7 before and after incorporation of the HER2-AuNPs was also obtained indicating a decrease in zeta potential with the incorporation of the nanoparticles. The outcome of this research will improve our fundamental understanding of the behavior of cancer cells and define optimal parameters of electrotherapy for clinical and drug delivery applications.
    Theranostics 01/2014; 4(9):919-30. · 7.81 Impact Factor
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    ABSTRACT: We have previously reported a local redox cycling-based electrochemical (LRC-EC) system for the incorporation of many electrochemical sensors into a small chip device. In the present study, a new type of LRC-EC chip device was fabricated for the detection of a droplet array. To detect electrochemically redox compounds in droplets, Pt pseudo-reference/counter electrodes were incorporated into the individual sensors of the LRC-EC chip device. Cyclic voltammetry for the LRC-EC chip device with internal Pt pseudo-reference electrodes indicated well-defined voltammograms based on redox cycling for the individual sensor points. The device was successfully applied to the addressable detection of alkaline phosphatase (ALP) activity of HeLa cells in single droplets on the sensor points. Therefore, the LRC-EC chip device is considered to be a useful device for the bioanalysis of droplet systems.
    Lab on a Chip 12/2013; · 5.70 Impact Factor
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    ABSTRACT: A novel measurement system to determine oxygen consumption rates via respiration in migrating Zebrafish (Danio rerio) has been developed. A signal equalization system was adapted to detect oxygen in a chamber with one fish, because typical electrochemical techniques cannot measure respiration activities for migrating organisms. A closed chamber was fabricated using a pipette tip attached to a Pt electrode and a columnar Vycor glass tip was used as the salt bridge. Pt electrode which was attached to the chamber with one zebrafish, and Ag electrode were immersed in 10 mM potassium iodide (KI), and both the electrodes were connected externally to form a galvanic cell. Pt and Ag electrodes act as the cathode and anode to reduce oxygen and oxidize silver, respectively, allowing the deposition of insoluble silver iodide (AgI). The AgI acts as the signal source accumulated on the Ag electrode by conversion of oxygen. The amount of AgI deposited on the Ag electrode was determined by cathodic stripping voltammetry. The presence of zebrafish or its embryo led to decrease in the stripping currents generated by a 10 min conversion of oxygen to AgI. The conversion of oxygen to AgI is disturbed by the migration of the zebrafish and allows the detection of different equalized signals corresponding to respiration activity. The oxygen consumption rates of the zebrafish and its embryo were estimated and determined to be ~4.1 and 2.4 pmol•s-1, respectively. The deposited AgI almost completely disappeared with a single stripping process. The signal equalization system provides a method to determine the respiration activities for migrating zebrafish and could be used to estimate environmental risk and for effective drug screening.
    Analytical Chemistry 12/2013; · 5.70 Impact Factor
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    ABSTRACT: In this study, we introduce the double-barrel carbon probe (DBCP)-a simple, affordable microring electrode-which enables the collection and analysis of single cells independent of cellular positioning. The target cells were punctured by utilizing an electric pulse between the two electrodes in DBCP, and the cellular lysates were collected by manual aspiration using the DBCP. The mRNA in the collected lysate was evaluated quantitatively using real-time PCR. The histograms of single-cell relative gene expression normalized to GAPDH were fit to a theoretical lognormal distribution. In the tissue culture model, we focused on angiogenesis to prove that multiple gene expression analysis was available. Finally, we applied DBCP for the embryonic stem (ES) cell-derived cardiomyocytes to substantiate the capability of the probe to collect cells, even from high-volume samples such as spheroids. This method achieves high sensitivity for mRNA at the single-cell level and is applicable in the analysis of various biological samples independent of cellular positioning.
    Analytical and Bioanalytical Chemistry 11/2013; · 3.66 Impact Factor
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    ABSTRACT: Amorphous hydrocarbon (aCH) material is receiving plenty of attention due to its possible wide application. However, one hurdle facing this application is that high temperature is required to express conductivity of aCH, e.g., post annealing or deposition at high temperature. To form a conductive aCH on a substrate controlled below room temperature, we have developed a neutral beam enhanced chemical vapor deposition (NBECVD) method to control a hydrocarbon molecular structure that has a large conjugated systemwith delocalized p electrons in film. For material gas, we prepared toluene. As a result, we obtained a highly conductive carbon on a Si substrate with �50 �C using only toluene by optimizing the state of disassociated material gas. From an evaluation of film structure, a polycyclic aromatic hydrocarbon molecular structure was grown and contained in film because NBECVD could avoid irradiating UV to the Si substrate during deposition. Thus, an excited large conjugated chain structure generated by toluene in plasma could be maintained and polymerized on the Si substrate. Furthermore, the conductive aCH film could work as electrode in solution by electrochemical examination. Additionally, we found that nitrogen doped into conductive aCH could increase the working current of an electrode.
    Carbon 10/2013; 67(February):635. · 5.87 Impact Factor
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    ABSTRACT: Alkaline phosphatase (ALP) is an enzyme commonly used as an undifferentiated maker of embryonic stem cells (ESCs). Although non-invasive ALP detection has long been desired for stem cell research and in cell transplantation therapy, little progress has been made in developing such techniques. In this study, we propose a non-invasive evaluation method for detecting ALP activity in mouse embryoid bodies (mEBs) using scanning electrochemical microscopy (SECM). SECM has several advantages, being non-invasive, non-labeled, quantitative, and highly sensitive. First, we found that SECM-based ALP evaluation permits the comparison of ALP activity among mEBs of different sizes by monitoring the p-aminophenol (PAP) production rate in aqueous solution containing p-aminophenylphosphate (PAPP) normal to the surface area of the each sample. Second, co-culture spheroids, consisting of mEB and MCF-7 cells for the core and the concentric outer layer, respectively, were prepared as model samples showing heterogeneous ALP activities. The overall PAP production rate dramatically declined in the presence of the MCF-7 cell outer layer, which blocked the mass transfer of PAPP to inner mEB. This result indicated that the SECM response mainly originated from ALP located at the surface of the cellular aggregate, including mEBs and co-culture spheroids. Third, taking advantage of the non-invasive nature of SECM, we examined the relevance of ALP activity and cardiomyocytes differentiation. Collectively, these results suggested that non-invasive SECM-based ALP activity normalized by the sample surface enables the selection of EBs with a higher potential to differentiate into cardiomyocytes, which can contribute toward various types of stem cell research.
    Analytical Chemistry 09/2013; · 5.70 Impact Factor
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    ABSTRACT: Mouse embryoid bodies (mEBs) were evaluated in detail on the basis of respiratory activity and high-throughput quantitative reverse transcription-PCR (RT-qPCR) analysis. The hanging drop culture method was applied to prepare various sizes of mEBs ranging from 100 to 250 μm in radius by causing the aggregation of embryonic cells. The respiratory activity of individual mEBs was noninvasively measured using scanning electrochemical microscopy in a cone-shaped microwell. For gene expression analysis, we used 48.48 Dynamic Array chips (Fluidigm) integrating microfluidic circuits, which allowed high-throughput qPCR analysis in parallel. The respiratory activity of the mEBs that were cultured for 1 to 6 days could predict the mRNA levels of undifferentiation and differentiation markers. However, the sizes of the mEBs could also predict the gene expression of the undifferentiation/differentiation markers because the radii of the mEBs increased by more than 2-fold after incubation in hanging drop culture for 6 days. Next, mEBs with identical sample sizes were evaluated for respiratory activity and gene expression. For mEBs cultured at 1500 cells per droplet for 3 days, the respiratory activity was negatively correlated with the mRNA levels of pluripotent markers such as Nanog and Sox2. Many differentiation markers were positively correlated with the respiratory activity. However, there was no significant difference in respiration activity between the beating and nonbeating samples on day 3. Finally, principal component analysis (PCA) confirmed the relationship between respiratory activity and the mRNA levels of undifferentiation/differentiation markers.
    Molecular BioSystems 08/2013; · 3.35 Impact Factor

Publication Stats

2k Citations
771.90 Total Impact Points

Institutions

  • 1979–2014
    • Tohoku University
      • • Graduate School of Environmental Studies
      • • Department of Bioengineering and Robotics
      • • Graduate School of Engineering
      • • Department of Applied Bioorganic Chemistry
      • • Department of Chemical Engineering
      • • School of Engineering
      Japan
  • 2009–2011
    • National Institute for Environmental Studies
      Tsukuba, Ibaraki, Japan
    • National Cheng Kung University
      • Institute of Biomedical Engineering (IBE)
      Tainan, Taiwan, Taiwan
  • 2009–2010
    • University of Hyogo
      • Graduate School of Material Science
      Kōbe-shi, Hyogo-ken, Japan
  • 2008
    • University of Southampton
      Southampton, England, United Kingdom
  • 2005
    • Kumamoto University
      Kumamoto, Kumamoto Prefecture, Japan
  • 2002
    • University of Glasgow
      • Division of Electronics and Electrical Engineering
      Glasgow, SCT, United Kingdom
  • 1987
    • University of Delaware
      • Department of Chemistry and Biochemistry
      Newark, DE, United States