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Manabu Tokeshi

Publications and source records attributed to Manabu Tokeshi.

At least 19 recordsLinked to original sources
PubMed2007

Quantum dots for single bio-molecule imaging.

The emerging nanomaterial, quantum dots or QDs, offers numerous potential applications in the biological area. As cell labeling probes, QDs become now an alternative of existing organic fluorescent dyes and fluorescent proteins. In this short review, we cover typical and successful applications of QDs as fluorescent probes in cell labeling and genomic diagnosis. As a future important application, biomolecular detection at a single molecule level utilizing QDs is also discussed.

Animals↗
PubMed2006

Pressure-driven flow control system for nanofluidic chemical process.

We developed a novel flow control system for a nanofluidic chemical process. Generally, flow control in nanochannels is difficult because of its high-pressure loss with very small volume flow rate. In our flow control method, liquid pressure in a microchannel connected to the nanochannels is regulated by utilizing a backpressure regulator. The flow control method was verified by using simple structured microchip, which included parallel nanochannels. We found that the observed flow rate was three times lower than the value expected from Hagen-Poiseuille's equation. That implied a size-dependent viscosity change in the nanochannels. Then, we demonstrated mixing of two different fluorescent solutions in a Y-shaped nanochannel and also a proton exchange reaction in the Y-shaped nanochannel. The flow control method will contribute to further integration of nanochemical systems.

Microfluidics↗
PubMed2006

Microchip-based liquid-liquid extraction for gas-chromatography analysis of amphetamine-type stimulants in urine.

A microchip-based liquid-liquid extraction for the gas chromatography analysis of urine for amphetamine-type stimulants has been developed. Partially modified microchannels with the capillarity restricted modification (CARM) method were employed for stabilizing the interface consisting of 1-chlorobutane and alkalized urine. Reliability of the microchip-based extraction was evaluated with respect to linearity, trueness and precision. As a practical demonstration, methoxyphenamine hydrochloride (50 mg) was administered to three healthy volunteers, and the concentration of methoxyphenamine in their urine was determined by both methods for comparison. This study showed the potential of pressure-driven microfluidics to contribute to the rapid automation analysis in forensic toxicology.

Adult↗
PubMed2006

Study of water properties in nanospace.

Here we report an anomalous behavior of water, especially its viscosity and hydrodynamic flow, in a nanometer-confined space. As a typical model of a nanometer-confined space, the nanopillar chip, which was developed for DNA size-based separation was used, and single-particle tracking (SPT) technique was applied to investigate water viscosity and hydrodynamic flow in the nanopillar chip. The diffusion coefficients of nanospheres were almost one-third of the theoretical value derived from the Stokes-Einstein equation. This result gave indirect proof that water viscosity in a nanometer-confined space is higher than in a bulk solution. In order to improve resolution and throughput of the nanopillar chip for DNA separation, these potential factors affecting performance should be seriously considered.

Journal Article↗
PubMed2006

Electrophoretic behavior of plasmid DNA in the presence of various intercalating dyes.

In the present study, the electrophoretic behavior of linear, supercoiled and nicked circular plasmid DNA in the presence of various intercalating dyes was characterized using pGL3 plasmid DNA as a model. The enzymatic digestion of pGL3 plasmid DNA with HindIIIwas monitored by capillary electrophoresis coupled with laser-induced fluorescence detection (CE-LIF). Nicked circular plasmid DNA was found to be relatively sensitive to enzymes, and was almost digested into the linear conformer after 10-min incubation, indicating that nicked circular plasmid DNA has little chance of targeting and entering the cell nucleus. Partly digested plasmid DNA containing only linear and supercoiled conformers can be used as a standard to confirm the migration order of plasmid DNA. In methylcellulose (MC) solution with YO-PRO-1 or YOYO-1, linear plasmid DNA eluted first, followed by supercoiled and nicked plasmid DNA, and nicked plasmid DNA eluted as a broad peak. With SYBR Green 1, nicked plasmid DNA eluted first as three sharp peaks, followed by linear and supercoiled plasmid DNA. The nuclear plasmid DNA from two transfected cell lines was successfully analyzed using the present procedure. Similar results were obtained with an analysis time of seconds using microchip electrophoresis with laser-induced fluorescence detection (mu-CE-LIF). To our knowledge, these results represent the first reported analysis of nuclear plasmid DNA from transfection cells by CE-LIF or mu-CE-LIF without pre-preparation, suggesting that the present procedure is a promising alternative method for evaluating transfection efficiency of DNA delivery systems.

3T3 Cells↗
PubMed2006

UV excitation thermal lens microscope for sensitive and nonlabeled detection of nonfluorescent molecules.

An ultrasensitive and nonlabeled detection method of nonfluorescent molecules on a microchip was developed by realizing a thermal lens microscope (TLM) with a 266-nm UV pulsed laser as an excitation light source (UV-TLM). Pulsed laser sources have advantages over continuous-wave laser sources in more compact size and better wavelength tuning, which are important for microchip-based analytical systems. Their disadvantage is difficulty in applying a lock-in amplifier due to the high (>10(4)) duty ratio of pulse oscillation. To overcome this problem, we realized a quasi-continuous-wave excitation by modulating the pulse trains at approximately 1 kHz and detecting the synchronous signal with a lock-in amplifier. The optimum pulse repetition frequency was obtained at 80 kHz, which was reasonable considering thermal equilibrium time. Furthermore, a permissible flow velocity in the range of 6.6-19.8 mm/s was found to avoid sensitivity decrease due to photochemical reactions and thermal energy dissipation. Under these conditions, we detected adenine aqueous solutions on a fused-silica microchip without labeling and obtained a sensitivity that was 350 times higher than that in a spectrophotometric method. The sensitivity was enough for detection on a microchip with an optical path length that was 2-3 orders shorter than that in conventional cuvettes. Finally, the UV-TLM method was applied to liquid chromatography detection. Fluorene and pyrene were separated in a microcolumn and detected in a capillary (50-microm inner diameter) with 150 times higher sensitivity than a spectrophotometric method. Our method provides highly sensitive and widely applicable detections for various analytical procedures and chemical syntheses on microchips.

Adenine↗
PubMed2006

Circular dichroism thermal lens microscope for sensitive chiral analysis on microchip.

A novel chiral detector, a circular-dichroism thermal lens microscope (CD-TLM), was developed to realize sensitive and selective detection of small volume chiral samples on a microchip. To realize chiral recognition on TLM, an excitation beam was phase-modulated at a frequency of 1.2 kHz, and left-circularly polarized light (LCPL) and right-circularly polarized light (RCPL) were generated. Then, the differential light absorption between LCPL and RCPL, which is the CD effect, was detected as thermal lens signal intensity and phase. As a standard sample, optically active tris(ethylenediamine)cobalt(III) [Co-(en)3]3+I3- aqueous solutions were used for performance evaluations. First, we verified the basic principle for selective chiral analysis by comparing the signals in intensity-modulation and phase-modulation modes of the excitation beam. Also, we found that the g-factor, which is significant for determining enantiomeric excess, agreed well with the value obtained by the CD spectrometer. The limit of detection (LOD) for enantiopure [Co-(en)3]3+I3- was 6.3 x 10(-5) M (1.9 x 10(-7) abs) for (-)-Co(en)3(3+), and the sensitivity in absorbance units was more than 250 times higher than that in a CD spectrophotometer. Finally, we demonstrated enantiomeric excess determination on a microchip. The LOD was 1.7% (8.5 x 10(-7) abs) for (-)-Co(en)3(3+) and at least one order superior to the LOD of a CD spectrometer. The applicability of CD-TLM for sensitive chiral analysis on a microchip was verified, and CD-TLM is expected to be promising for microchip-based chiral synthesis and analysis systems.

Journal Article↗
PubMed2006

Rapid multiplexing and simultaneous detection of human spermatogenetic failure with a 12 lane microchip electrophoresis system.

For the amplification and ultrafast separation of the genetic markers and DNA sequences that are related to human male infertility, a multiplex PCR for amplifying three DNA sequence-tagged sites (STS) located on the human Y chromosome with possible roles in the spermatogenesis process has been designed and applied followed by separation on a microchip. First, the optimum T(m) degree for the three DNA markers was optimized and determined experimentally, and the three DNA STS were amplified. These three DNA markers were then separated on a 12-lane microchip electrophoresis system, which can analyze the DNA markers on 12 channels simultaneously. The combination of these two technologies, multiplex PCR and microchip electrophoresis, allows the analysis of 36 DNA markers (12x3) within only 180 s.

Base Sequence↗
PubMed2006

Low viscous separation media for genomics and proteomics analysis on microchip electrophoresis system.

Microchip electrophoresis has widely grown during the past few years, and it has showed a significant result as a strong separation tool for genomic as well as proteomic researches. To enhance and expand the role of microchip electrophoresis, several studies have been proposed especially for the low viscous separation media, which is an important factor for the success of microchip with its narrow separation channels. In this paper we show an overview for the done researches in the field of low viscous media developed for the use in microchip electrophoresis. For genomic separation studies polyhydroxy additives have been used enhance the separation of DNA at low polymer concentration of HPMC (Hydroxypropylmethyl cellulose) which could keep the viscosity low. Mixtures of poly(ethylene oxide) as well as Hydroxyporpyl cellulose have been successfully introduced for chip separation. Furthermore high molecular mass polyacrylamides at low concentrations have been studied for DNA separation. A mixture of polymer nanoparticle with conventional polymers could show a better resolution for DNA at low concentration of the polymer. For the proteomic field isoelectric focusing on chip has been well overviewed since it is the most viscous separation media which is well used for the protein separation. The different types of isoelectric focusing such as the ampholyte-free type, the thermal type as well as the ampholyte-depended type have been introduced in this paper. Isoelectric focusing on chip with its combination with sodium dodecyl sulfate (SDS) page or free solution could give a better separation. Several application for this low viscous separation medias for either genomic or proteomic could clearly show the importance of this field.

Animals↗
PubMed2006

Liquid filling method for nanofluidic channels utilizing the high solubility of CO2.

We developed a fabrication method and a liquid filling method for a nano chemical reactor that used Y-shaped nanochannels specially designed for mixing and reacting. In order to reduce the pressure loss and to utilize the characteristics of the nanochannel, inlet microchannels were fabricated just beside the nanochannels. We investigated an initial liquid filling method into the nanochannels that ensured there were no air bubbles that could cause a flow stack due to the capillary pressure. In our method, the micro- and nanochannels were filled with carbon dioxide and any remaining air during the initial liquid introduction was dissolved utilizing the high solubility of carbon dioxide. We propose that chemical reactions in nanospaces can be realized by utilizing these fabrication and liquid introduction techniques.

Journal Article↗
PubMed2006

Quantitative detection and fixation of single and multiple gold nanoparticles on a microfluidic chip by thermal lens microscope.

A detection and fixation method of single and multiple gold nanoparticles on the wall of a microfluidic channel is demonstrated. A thermal lens microscope (TLM) with continuous-wave excitation (wavelength, 532 nm) and probe (wavelength, 670 nm) laser beams was used to realize the sensitive detection of heat generated by light absorption of individual gold nanoparticles (50 nm in diameter); fixation of the individual nanoparticles was realized simultaneously. The fixation mechanism was investigated and attributed to an absorption-based optical force. In addition to single nanoparticle detection, multiple-nanoparticle detection and fixation was demonstrated. An acceleration of fixation was observed when the number of fixed particles was increased. TLM is expected to be a powerful tool for both the quantitative detection and precise fixation of individual nanoparticles.

Journal Article↗
PubMed2005

Rapid analysis of oligosaccharides derived from glycoproteins by microchip electrophoresis.

A novel method for fast profiling of complex oligosaccharides released from glycoproteins based on microchip electrophoresis (mu-CE) is presented here. The characterization of separation conditions, i.e., the composition, concentration and pH of running buffer as well as the applied voltage, has been performed using maltose (G2), cellobiose ( G2'), maltriose (G3) and panose (G3') as oligosaccharide isomer models. In mu-CE, much better separation of oligosaccharide isomers and oligosaccharide ladder was obtained in phosphate buffer than in borate buffer over a wide pH range. Under optimal conditions, high-performance separation of the N-linked complex oligosaccharides released from ribonuclease B, fetuin, alpha1-acid glycoprotein (AGP) and IgG was achieved using polymethylmethacrylate (PMMA) microchips with an effective separation channel of 30 mm. These results represent the first reported analysis of the N-linked oligosaccharides derived from glycoproteins by mu-CE, indicating that the present mu-CE-based method is a promising alternative for characterization of the N-linked oligosaccharides in glycoproteins.

Carbohydrate Sequence↗
PubMed2005

Miniaturized thermal lens and fluorescence detection system for microchemical chips.

We have developed a miniaturized two-way detection system using thermal lens and fluorescence spectroscopies for microchip chemistry. The system was composed of laser diode (LD) modules, fiber-based optics combined with a gradient index lens, and miniaturized detection units for thermal lens and fluorescence signals. The detection limits in the thermal lens and fluorescence spectroscopies were 6.3 x 10(-9)M for Ni(II) phthalocyanine tetrasulfonic acid and 3.0 x 10(-9)M for cy5, respectively. The performance of the system with the miniaturized thermal lens was equivalent to that of a conventional thermal lens microscope. The fluorescence sensitivity was comparable to sensitivities offered by conventional miniaturized systems.

Miniaturization↗
PubMed2005

Monitoring of intercellular messengers released from neuron networks cultured in a microchip.

A cellular biochemistry analysis system was integrated on a quartz glass microchip with a microchamber for cell culture followed by a microchannel for detecting with a thermal lens microscope (TLM). Nerve cells from rat hippocampus were successfully cultured to form neural networks in the microchip. An aqueous solution of glutamate, which is known as a neurotransmitter, was introduced to stimulate the cultured neuron to release a retrograde messenger, arachidonate which is considered to be critical for neuronal plasticity, especially for long-term potentiation (LTP). After the introduction, the solution that flowed through the culture chamber was analyzed using the UV-TLM (excitation wavelength, 244 nm). The measured signal intensity was dependent on glutamate solution concentration, and the neurons were considered to release the retrograde messenger according to the glutamate concentration. This system is suitable for time-course monitoring of ultra trace amounts of chemicals released from very small amount of cultured cells.

Animals↗
PubMed2005

Development of a microchip-based bioassay system using cultured cells.

We developed a novel bioassay system using a glass microchip and cultured cells. A microchamber for cell culture and microchannels for reactions and detection were fabricated on a Pyrex glass substrate by photolithography and wet etching techniques. Cell culture, chemical and enzymatic reactions, and detection were integrated into the microchip. To keep different temperatures locally in three areas of the microchip, we designed and fabricated a temperature control device. Nitric oxide released from macrophage-like cells stimulated by lipopolysaccharide was successfully monitored with the microchip, the temperature control device, and a thermal lens microscope. The total assay time was reduced from 24 to 4 h, and detection limit of NO was improved from 1 x 10(-6) to 7 x 10(-8) M compared with conventional methods. Moreover, the system could monitor a time course of the release, which is difficult to measure by conventional batch methods. We conclude that this system is promising for a rapid bioassay system with very small consumption of cells.

Animals↗
PubMed2005

Surface modification method of microchannels for gas-liquid two-phase flow in microchips.

A capillarity restricted modification method for microchannel surfaces was developed for gas--liquid microchemical operations in microchips. In this method, a microstructure combining shallow and deep microchannels and the principle of capillarity were utilized for chemical modification of a restricted area of a microchannel. A hydrophobic--hydrophilic patterning in microchannels was prepared as an example for guiding gas and liquid flows along the respective microchannels. Validity of the patterning was confirmed by measuring aqueous flow leak pressure from the hydrophilic microchannel to the hydrophobic one. The leak pressure of 7.7-1.1 kPa agreed well with that predicted theoretically from the Young-Laplace equation for the microchannel depth of 8.6-39 microm. In an experiment to demonstrate usefulness and effectiveness of the method, an air bubble was first introduced into the hydrophilic microchannel and purged from the hydrophobic-hydrophilic patterned microchannels. Next, the patterning structure was applied to remove dissolved oxygen by contacting the aqueous flow with a nitrogen flow. The concentration of dissolved oxygen decreased with contact time, and its time course agreed well with numerical simulation. These demonstrations showed that the proposed patterning method can be used in general microfluidic gas-liquid operations.

Biomechanical Phenomena↗
PubMed2005

Thermal lens micro optical systems.

This paper describes two types of miniaturized thermal lens optical systems that use optical fibers, SELFOC microlenses and light sources. The first system consists of a compact diode pumped solid-state laser (532 nm) as an excitation light source, a laser diode (635 nm) as a probe light source, an acoustoptic modulator as an excitation light modulator, fiber-based and conventional optics, and a detection system that combines a pinhole, an interference filter, and a photodiode. The second system consists of two laser diodes as the excitation (658 nm) and probe (780 nm) light sources, fiber-based optics, and the same detection system as the first one. The performance of the two systems was evaluated by the limit of detection (LOD) using standard solutions of sunset yellow (SY) and nickel(II) phthalocyaninetetrasulfonic acid tetrasodium salt (NiP). The LODs of the first system for SY and second system for NiP were calculated to be 3.7 x 10(-8) (1.7 x 10(-6) AU) and 7.7 x 10(-9) M (3.4 x10(-6) AU), respectively. These results were consistent with the expected values obtained from photothermal parameters.

Journal Article↗