研究者詳細 - 金田　隆

論文 - 金田　隆

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Determination of polyamines in Arabidopsis thaliana by capillary electrophoresis using salicylaldehyde-5-sulfonate as a derivatizing reagent 査読

Genki Inoue, Takashi Kaneta, Toshio Takayanagi, Junichi Kakehi, Hiroyasu Motose, Taku Takahashi

ANALYTICAL METHODS 5 ( 11 ) 2854 - 2859 2013年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ROYAL SOC CHEMISTRY

Herein, we report a novel method for the determination of polyamines in a sample extracted from Arabidopsis thaliana by capillary electrophoresis (CE) using salicylaldehyde-5-sulfonate (SAS) as a derivatizing reagent. An aldehyde group of SAS forms a Schiff base with amino groups of aliphatic polyamines, resulting in an anionic species with an absorption band in the ultraviolet region. The derivatization method was straightforward since the derivatives were formed by mixing a sample with the derivatizing reagent at a neutral pH. In addition, the negative charges induced by SAS led to a high resolution with a short analysis time. This method permitted the separation of five polyamines, which play important roles in plants. However, further improvement in sensitivity was needed for the determination of the polyamines in plant samples. Therefore, the CE method was coupled with solid-phase extraction (SPE) using an ion-pairing formation with sodium dodecyl benzene sulfonate. The SPE method improved the concentration limits of detection to sub-mu M levels, which corresponded with a 10-fold enhancement. The calibration curves for cadaverine, putrescine, and spermidine were linear with concentrations that ranged from 1 to 20 mu M and correlation coefficients (R-2) were greater than 0.998. The proposed method was applied to the determination of spermidine in a plant sample, Arabidopsis thaliana.

DOI： 10.1039/c3ay26360f

Sheath-flow electrochemical detection of amino acids with a copper wire electrode in capillary electrophoresis 査読

Junji Inoue, Takashi Kaneta, Totaro Imasaka

ELECTROPHORESIS 33 ( 17 ) 2743 - 2747 2012年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

Here, we report the detection of native amino acids using a sheath-flow electrochemical detector with a working electrode made of copper wire. A separation capillary that was inserted into a platinum tube in the detector acted as a grounded electrode for electrophoresis and as a flow channel for sheath liquid. Sheath liquid flowed outside the capillary to support the transport of the separated analytes to the working electrode for electrochemical detection. The copper wire electrode was aligned at the outlet of the capillary in a wall-jet configuration. Amino acids injected into the capillary were separated following elution from the end of the capillary and detection by the copper electrode. Three kinds of copper electrodes with different diameters50, 125, and 300 mu mwere examined to investigate the effect of the electrode diameter on sensitivity. The peak widths of the analytes were independent of the diameter of the working electrode, while the 300-mu m electrode led to a decrease in the signal-to-noise ratio compared with the 50- and 125-mu m electrodes, which showed no significant difference. The flow rate of the sheath liquid was also varied to optimize the detection conditions. The limits of detection for amino acids ranged from 4.4 to 27 mu M under optimal conditions.

DOI： 10.1002/elps.201200059

Determination of Arsenic(III) by Sequential Injection/ Anodic Stripping Voltammetry (SI/ASV) Using In-situ Thin Film-Modified Screen-Printed Carbon Electrode (SPCE) 査読

Eakkasit Punrat, Suchada Chuanuwatanakul, Orawan Chailapakul, Toshio Takayanagi, Takashi Kaneta, Shoji Motomizu

Journal of Flow Injection Analysis 29 ( 1 ) 11 - 16 2012年6月

Band broadening of DNA fragments isolated by polyacrylamide gel electrophoresis in capillary electrophoresis 査読

Takashi Kaneta, Takehito Ogura, Shuhei Yamato, Totaro Imasaka

JOURNAL OF SEPARATION SCIENCE 35 ( 3 ) 431 - 435 2012年2月

担当区分：筆頭著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY PERIODICALS, INC

Polyacrylamide gel electrophoresis (PAGE) is used frequently for isolation and purification of DNA fragments. In the present study, DNA fragments extracted from polyacrylamide gels showed significant band broadening in capillary electrophoresis (CE). A pHY300PLK (a shuttle vector functioning in Escherichia coli and Bacillus subtilis) marker, which contained nine fragments ranging from 80 to 4870?bp, was separated by PAGE, and each fragment was isolated by phenol/chloroform extraction and ethanol precipitation. After extraction from the polyacrylamide gel, the peaks of the isolated DNA fragments exhibited band broadening in CE, where a linear poly(ethylene oxide) was used as a sieving matrix. The theoretical plate numbers of the DNA fragments contained in the pHY300PLK marker were >106 for all the fragments before extraction. However, the DNA fragments extracted from the polyacrylamide gel showed decreased theoretical plate numbers (520 times smaller). The degradation of the theoretical plate number was significant for middle sizes of the DNA fragments ranging from 489 to 1360?bp, whereas the largest and smallest fragments (80 and 4870?bp) had no obvious influence. The band broadening was attributed to contamination of the DNA fragments by polyacrylamide fibers during the separation and extraction process.

DOI： 10.1002/jssc.201100909

Micellar electrokinetic chromatographic analysis for in vitro accumulation of anthracyclines enhanced by inhibitors of cell membrane transporter-proteins in cancer cells 査読

Julius Mbuna, Takashi Kaneta, Totaro Imasaka

BIOMEDICAL CHROMATOGRAPHY 25 ( 10 ) 1168 - 1174 2011年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY

Cell membrane transporter-proteins have been partly implicated in lowering the accumulation of drugs in cancer cells, leading to multidrug resistance (MDR). Two cancer cell lines, A549 and RDES, were continuously exposed to subclinical concentration (250 nM) of anthracyclines and micellar electrokinetic chromatography was used to investigate their in vitro accumulation after treatment with inhibitors of membrane transporter-proteins. The four anthracylines [doxorubicin (DOX), epirubicin (EPI), daunorubicin (DNR), and idarubicin (IDA)] were separated within a short analysis time of less than 15 min in borate buffer (80 mM, pH 9.22) containing sodium taurodeoxycholate (35 mM), 2-hydroxypropyl-.-cyclodextrin (3.5% wt/v), and sodium dodecylsulfate (20 mM). Laser-induced fluorescence was used for detection of the anthracyclines. Three inhibitors, verapamil, cyclosporine A and probenecid, were examined by adding each inhibitor independently or two inhibitors simultaneously to the culture medium. It was found that independent use of each inhibitor leads to more efficient accumulation than combined use of verapamil and probenecid. In addition, the results show that effect of inhibitors on the accumulation of anthracyclines depended on type of cell: in RDES, inhibitors enhanced accumulation of all four anthracyclines, while in A549, inhibitors showed different accumulation behavior for each anthracycline. Generally higher accumulation of anthracyclines was observed in RDES cells than A549, as evidenced by dead cells (7-16%) after 24 h of continuous exposure to subclinical concentration. Copyright (C) 2011 John Wiley & Sons, Ltd.

DOI： 10.1002/bmc.1589

Rapid determination of multidrug resistance-associated protein in cancer cells by capillary electrophoresis immunoassay 査読

Julius Mbuna, Takashi Kaneta, Totaro Imasaka

JOURNAL OF CHROMATOGRAPHY A 1218 ( 25 ) 3923 - 3927 2011年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER

The adenosine triphosphate (ATP) binding-cassette (ABC) transporters are a superfamily of cellular proteins that have been partly implicated as a cause of multidrug resistance (MDR) in cancer cells. The ABC superfamily consists of P-glycoprotein, multidrug resistance-associated proteins (MRP) and breast cancer-related proteins, of which MRP is of particular interest because of its ability to efflux a broader range of substrates. Since MRP1 is the most prominent member of the MRP family, a simple technique is needed for its quantification. We developed a simple, fast (total analysis time of 3 h) capillary electrophoresis immunoassay (CEIA) for the quantification of MRP1 in cancer cells. MRP1 antibody was labeled with fluorescein isothiocyanate. The labeled antibody was incubated with the cell lysate for a fixed interval (1 h), after which the cell lysate mixture was directly injected into the capillary to separate the complex of MRP1 and its antibody from free antibody. The noncompetitive CEIA method had a limit of detection of 0.2 nM and a good linear range (1.7-14.9 x 10(4) cells), and was fairly reproducible (RSD < 10%). The results showed that two cell lines. A549 and RDES, expressed MRP1 in the absence of doxorubicin (DOX), with A549 registering a higher expression. Compared to DOX-free cancer cells, there was an acceleration of MRP1 expression during the 12 h-exposure to DOX, after which the level of expression remained nearly constant as the intracellular accumulation of DOX decreased. The results obtained in this work indicate that the developed CEIA method is useful for relative quantification of MRPs in cancer cells. (C) 2011 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.chroma.2011.04.046

Analysis of proteins in biological samples by capillary sieving electrophoresis with postcolumn derivatization/laser-induced fluorescence detection 査読

Takashi Kaneta, Takehito Ogura, Totaro Imasaka

ELECTROPHORESIS 32 ( 9 ) 1061 - 1067 2011年4月

担当区分：筆頭著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

Previously, we have demonstrated postcolumn derivatization of proteins separated by capillary sieving electrophoresis (CSE), in which naphthalene-2,3-dicarbaldehyde was employed as a fluorogenic labeling reagent. Standard proteins separated by CSE were reacted with naphthalene-2,3-dicarbaldehyde in the presence of 2-mercaptoethanol (2-ME) which plays a role of a reducing agent in the derivatization reaction. To improve the sensitivity, we attempted the use of ethanethiol instead of 2-ME. Ethanethiol showed 1.4- to 4.5-fold lower limits of detection for proteins than 2-ME. Furthermore, we found that 8-aminopyrene-1,3,6-trisulfonate (APTS) is a good marker for relative electrophoretic mobilities of proteins in CSE. Since APTS is a fluorescent and trivalent anion, it generates strong fluorescence and migrates faster than any of the proteins. Therefore, we employed APTS as a marker to obtain the relative electrophoretic mobilities of proteins. The present method was applied to the analyses of proteins in biological samples. Human Ewing's family tumor cell line 'RDES' was used as a sample. The cultured cells were lysed with a buffer containing Tris-HCl, NaCl, sodium dodecyl sulfate, and 2-ME. After denaturation, the lysate was directly introduced into the capillary. Several peaks, which would correspond to proteins with molecular mass ranging from 10 to 93 kDa, were found in the cell lysate. In addition, we measured a milk sample by the CSE with postcolumn derivatization. The electropherogram showed five major peaks which corresponded to alpha-lactalbumin, beta-lactoglobulin, kappa-casein, bovine serum albumin, and mixture of alpha- and beta-casein.

DOI： 10.1002/elps.201000488

Applications of Hadamard transform-gas chromatography/mass spectrometry to online detection of exhaled breath after drinking or smoking 査読

Yuan-Kai Cheng, Cheng-Huang Lin, Takashi Kaneta, Totaro Imasaka

JOURNAL OF CHROMATOGRAPHY A 1217 ( 32 ) 5274 - 5278 2010年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE BV

A Hadamard transform-gas chromatography/mass spectrometry (HT-GC/MS) technique was employed for the online detection of ethanol or toluene in exhaled breath after drinking or smoking, respectively. Exhaled breath samples, collected from volunteers, were directly injected into the GC inlet by a Hadamard-injector without any pretreatment. In the case of breath from a drinker, using a conventional single injection, a small ion peak (corresponding to similar to 0.1 ng of ethanol), the intensity of which was approximately equal to or less than the limit of detection. When the HT technique was applied, the signal-to-noise (S/N) ratio was dramatically improved. Furthermore, in the case of breath from a smoker, using conventional injection, a weak ion peak (corresponding to similar to 0.7 pg of toluene) was marginally detected. However, the HT technique led to an improvement in the S/N ratio, with the peak corresponding to the limit of detection. In both cases, the HT technique permitted specific components in exhaled breath to be determined, without the need for any extraction procedures. (C) 2010 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.chroma.2010.06.034

Measurement of intracellular accumulation of anthracyclines in cancerous cells by direct injection of cell lysate in MEKC/LIF detection 査読

Julius Mbuna, Takashi Kaneta, Totaro Imasaka

ELECTROPHORESIS 31 ( 8 ) 1396 - 1404 2010年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY

Anthracyclines are chemotherapeutic drugs that are broadly used in the treatment of various types of solid cancers and leukemia. Herein, we report on a novel analytical method for intracellular accumulation of anthracyclines using MEKC/LIF detection. An aqueous separation system permitted the injection of cell lysates directly into the capillary. The MEKC migrating solution was made up of borate buffer at pH 9.22 containing sodium taurodeoxycholate, (2-hydroxypropyl)-gamma-CD, and SDS. The anthracyclines, Doxorubicin (DOX) and epirubicin (EPI) were detected by LIF using a Nd:YAG laser (532 nm) or an argon ion laser (488 nm) for excitation. Two cell lines, human humerus tumor cells (RDES) and human lung tumor cells (A549), were treated with a mixture of the two anthracyclines for fixed periods of time, and then intracellular concentrations were determined by injecting cell lysates directly. Recovery values of 96.0-100.8% were obtained for DOX and EPI. Reproducibility quantified by RSD was less than 3.9% intraday and 6.7% interday at concentrations ranging between 50 and 500 nM. The uptake of EPI was found to be slightly less than that of DOX for A549, but higher levels of EPI were observed in RDES. Intracellular accumulation of anthracyclines was greater in RDES than in A549, but both types of cells excreted anthracyclines after 12 h. These results indicate that MEKC with an aqueous medium is useful for investigating intracellular uptake and accumulation of drugs, since cell lysates can be used directly with no pretreatment such as deproteination or solvent extraction of analytes.

DOI： 10.1002/elps.200900659

Application of Hadamard transform to gas chromatography/nonresonant multiphoton ionization/time-of-flight mass spectrometry 査読

Chao-Chiang Cheng, Hung-Wei Chang, Tomohiro Uchimura, Totaro Imasaka, Takashi Kaneta, Cheng-Huang Lin

JOURNAL OF SEPARATION SCIENCE 33 ( 4-5 ) 626 - 630 2010年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-V C H VERLAG GMBH

The technique of Hadamard transform was successfully coupled with GC/nonresonant multiphoton ionization/TOFMS, for the first time. 1,4-Dichlorobenzene and the fourth harmonic generation (266 nm) of a Nd:YAG laser were employed as a model sample and an ionization laser, respectively. A Hadamard-injector coupled with a capillary-based supersonic jet nozzle (capillary-injector) was also developed in this study. The Hadamard-injector was used to obtain the chromatogram, which was encoded by successive sample introduction based on Hadamard codes, and the capillary-injector was used for injection of GC-elutes into TOFMS. Compared with a conventional single injection method, the S/N ratios were substantially improved after inverse Hadamard transformation of the encoded chromatogram. Under optimized conditions, when Hadamard matrices of 103 and 255 were used, the S/N ratios of the signals for 1,4-dichlorobenzene (concentration level, 4 mu g/1 mL ACN) were substantially improved to 4.1- and 6.6-fold, respectively, and those improvements are in good agreement with those obtained by theory (5.1- and 8.0-fold).

DOI： 10.1002/jssc.200900662

Design and application of Hadamard-injectors coupled with gas and supercritical fluid sample collection systems in Hadamard transform-gas chromatography/mass spectrometry 査読

Zenzen Fan, Chien-Hung Lin, Hung-Wei Chang, Takashi Kaneta, Cheng-Huang Lin

JOURNAL OF CHROMATOGRAPHY A 1217 ( 5 ) 755 - 760 2010年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE BV

A novel Hadamard transform-gas chromatography/mass spectrometry (HT-GC/MS) system equipped with on-line sample collection systems is described. A Hadamard-injector was successfully designed and then coupled with an on-line adsorption/desorption system for detecting volatile organic compounds (VOCs) and a supercritical fluid extraction (SFE) system, respectively, by HT-GC/MS. Six VOCs and three pesticides were used as model compounds. In the former case, an activated-charcoal trap was used to trap VOCs from the indoor air. After 10 L of indoor air had passed through the trap, the condensed components were heated and simultaneously injected into the GC column through the Hadamard-injector, based on Hadamard codes. In a second experiment, a sample of rice was spiked with three types of pesticides and the sample then extracted using a commercially available supercritical fluid extractor. After extraction, the extracted components were transferred to a holding tank and simultaneously injected into the GC column also using the Hadamard-injector. The findings show that, in both cases, the combination of on-line sample collection methods and the use of the Hadamard transform resulted in improved sensitivity and detection. Compared to the single injection used in most GC/MS systems, the signal-to-noise (S/N) ratios were substantially improved after inverse Hadamard transformation of the encoded chromatogram. (C) 2009 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.chroma.2009.12.007

Transformation Techniques for Capillary and Microchip Electrophoresis 査読

Takashi Kaneta

Chemometric Methods in Capillary Electrophoresis 389 - 406 2009年11月

Postcolumn derivatization of proteins in capillary sieving electrophoresis/laser-induced fluorescence detection 査読

Takashi Kaneta, Daisuke Yamamoto, Totaro Imasaka

ELECTROPHORESIS 30 ( 21 ) 3780 - 3785 2009年11月

担当区分：筆頭著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-V C H VERLAG GMBH

The separation methods for proteins with high resolution and sensitivity are absolutely important in the field of biological sciences Capillary sieving electrophoresis (CSE) is an excellent separation technique for DNA and proteins with high resolution, while LIF permits the most sensitive detection in CSE. Therefore, proteins have to be labeled with fluorescent or fluorogenic reagent to produce fluorescent derivatives. Both precolumn and oncolumn derivatization have been employed for the labeling of proteins in CSE. However, there is no report on the postcolumn derivatization due to the limitation in the use of a standard migration buffer, despite it being a promising method for sensitive detection of proteins. Here, we show a novel postcolumn derivatization method for protein separation by CSE, using a tertiary amine as a buffer component in the running buffer. Tris, which is commonly used as a base in CSE separation buffers, was substituted by tertiary amines, 2-(diethylamino) ethanol and triethanolamine. A buffer solution containing 2-(diethylamino)ethanol or triethanolamine can be used for the CSE separation followed by the postcolumn derivatization. of proteins, since both reagents are unreactive toward a fluorogenic labeling reagent, naphthalene-2,3-dicarbaldehyde. Thus, LIF detection using the postcolumn derivatization. permits significant reduction in the LOD (by a factor of 2.4-28) of proteins, compared with conventional absorbance detection.

DOI： 10.1002/elps.200900314

Hadamard transform microchip electrophoresis combined with laser-induced fluorescence detection using a compact neodymium-doped yttrium aluminum garnet laser emitting at 532 nm 査読

Kazuki Hata, Takashi Kaneta, Totaro Imasaka

JOURNAL OF APPLIED PHYSICS 105 ( 10 ) 102018 2009年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER INST PHYSICS

Hadamard transform electrophoresis combined with laser-induced fluorescence (LIF) detection on a microchip was demonstrated. A compact, diode-pumped neodymium-doped yttrium aluminum garnet laser was employed as the light source for LIF detection. The analytical conditions were optimized using rhodamine B as the analyte. Under optimal conditions, the signal-to-noise ratio (S/N) of the analyte was improved by a factor of 7.5 by means of Hadamard transformation based on a 255-order cyclic S matrix. Additionally, the relationship between fluorescence intensity and analyte concentration was linear with a correlation coefficient of 0.993 in the inverse Hadamard transformed data at the concentration range from 25 to 100 pM. The results indicate that the present method is applicable to quantitative analysis at the concentration lower than the concentration limit of detection in a conventional method. The concentration limit of detection was similar to 25 pM (the relative standard deviation of the peak height was 5.2%). The present technique was successfully applied to the separation of a mixture containing 1.9 nM phenylalanine and 1.9 nM glutamic acid labeled with rhodamine B isothiocyanate. The S/Ns of the analyte peaks were improved up to similar to 10 in the inverse Hadamard transformed data derived from a 127-order cyclic S matrix, while neither peak was lower than the limit of detection (S/N < 3) in conventional microchip electrophoresis by a single injection.

DOI： 10.1063/1.3116121

Determination of amino acids in urine by cyclodextrin-modified capillary electrophoresis - Laser-induced fluorescence detection 査読

Takashi Kaneta, Hiromasa Maeda, Mari Miyazaki, Rina Miyake, Hirotomo Izaki, Yuji Sakoda, Shohei Kinoshita, Totaro Imasaka

JOURNAL OF CHROMATOGRAPHIC SCIENCE 46 ( 8 ) 712 - 716 2008年9月

Applications of Hadamard transform to gas chromatography mass spectrometry and liquid chromatography mass spectrometry 査読

Cheng-Huang Lin, Takashi Kaneta, Hung-Ming Chen, Wen-Xiong Chen, Hung-Wei Chang, Ju-Tsung Liu

ANALYTICAL CHEMISTRY 80 ( 15 ) 5755 - 5759 2008年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER CHEMICAL SOC

Successful application of the Hadamard transform (HI) technique to gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/mass spectrometry (LC/MS) is described. Novel sample injection devices were developed to achieve multiple sample injections in both GC and LC instruments. Air pressure was controlled by an electromagnetic valve in GC, while a syringe pump and Tee connector were employed for the injection device in LC. Two well-known, abused drugs, 3,4-methylenedioxy-N-methylamphetamine (MDMA) and N,N-dimethyltryptamine (DMT), were employed as model samples. Both of the injection devices permitted precise successive injections, resulting in clearly modulated chromatograms encoded by Hadamard matrices. After inverse Hadamard transformation of the encoded chromatogram, the signal-to-noise (S/N) ratios of the signals were substantially improved compared with those expected from theoretical values. ne S/N ratios were enhanced similar to 10-fold in HT-GC/MS and 6.8 in HT-LC/MS, using the matrices of 1023 and 511, respectively. The HT-GC/MS was successfully applied to the determination of MDMA in the urine sample of a suspect.

DOI： 10.1021/ac800201r

Postcolumn reactor using a laser-drilled capillary for light-emitting diode-induced fluorescence detection in CE 査読

Daisuke Yamamoto, Takashi Kaneta, Totaro Imasaka

ELECTROPHORESIS 28 ( 22 ) 4143 - 4149 2007年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-V C H VERLAG GMBH

This study investigated a novel postcolumn reactor for fluorescence detection in CE. A laser-drilled capillary, with an aperture made by laser ablation, was used for mixing derivatization reagents with the analytes separated by CZE. The derivatization reagents, o-phthaldialdehyde (OPA), and 2-mercaptoethanol, were introduced into the capillary through the aperture and reacted with the analytes after CZE separation. High voltages were applied to both the inlet reservoir and the reservoir filled with the derivatization reagents. Thus, the flow rate of the derivatization reagents was controlled by the electric potential that was applied to the reservoir of the derivatization reagents. A UV light-emitting diode was used as an excitation light source for the fluorescence detection of OPA derivatives. A commercially available tee connector was compared with the laser-drilled capillary. The results implied that the dead volume of the laser-drilled capillary was less than that of the tee connector since the laser-drilled capillary suppressed band broadening more efficiently. The LODs for amino acids were determined to be similar to 5 mu M. The method was applied to the determination of amino acids in a Japanese beverage.

DOI： 10.1002/elps.200700274

Highly sensitive analysis based on laser-induced fluorescence Detection/Capillary electrophoresis 査読

Takashi Kaneta

BUNSEKI KAGAKU 56 ( 10 ) 841 - 849 2007年10月

担当区分：筆頭著者記述言語：日本語掲載種別：研究論文（学術雑誌）出版者・発行元：JAPAN SOC ANALYTICAL CHEMISTRY

Laser-induced fluorescence detection using diode lasers as excitation light sources has been applied to the highly sensitive analysis of polyaromatic hydrocarbons, amino acids, DNA, and proteins. Diode lasers emitting at the near-infrared to red region were employed for direct and indirect fluorescence detections. In addition, to enhance the sensitivity in capillary electrophoresis, Hadamard transform capillary electrophoresis has been developed. The method includes multiple injection, followed by an inverse Hadamard transformation. Optically gated injection, electrokinetic injection using a microchip, and electrokinetic injection using a laser-fabricated capillary have been developed for achieving multiple sample injection. These methods have permitted multiple sample injection in capillary electrophoresis. The results obtained by Hadamard transform capillary electrophoresis showed a significant improvement in the S/N.

DOI： 10.2116/bunsekikagaku.56.841

On-column capture of a specific protein separated by SDS-CGE using an immunological reaction on magnetic beads 査読

Takashi Kaneta, Masanori Takahashi, Totaro Imasaka

ELECTROPHORESIS 28 ( 13 ) 2262 - 2266 2007年7月

担当区分：筆頭著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY

The on-column capture of a specific protein using magnetic beads was applied to SDS-CGE. In a preliminary study, an immunological reaction in the presence of SDS, using a batch method, was attempted. Carbonic anhydrase (CA), alpha-lactalbumin (LA), and HSA were denatured by heating in the presence of SDS and 2-mercaptoethanol, and then reacted with anti-CA that had been immobilized on magnetic beads. Not only native CA, but also the denatured CA reacted with anti-CA, even in the presence of SDS. Therefore, the on-column capture of denatured CA separated by SDS-CGE was attempted using a two-point detection technique. A mixture of proteins, containing LA, CA, and HSA, were separated by SDS-CGE according to their M-Gamma. The CA was then specifically captured on anti-CA-immobilized magnetic beads, which were packed between two detection windows in the capillary column, during the electrophoresis. The results show that the technique leads to information similar to that obtained by Western blotting, i.e., a protein can be identified by its M-Gamma and reaction with its antibody.

DOI： 10.1002/elps.200600726

Application of Hadamard transformation to MEKC 査読

Kazuki Hata, Takashi Kaneta, Totaro Imasaka

ELECTROPHORESIS 28 ( 3 ) 328 - 334 2007年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-V C H VERLAG GMBH

The Hadamard transform (HT) technique, which permits the S/N in CE to be improved, was applied to MEKC. Multiple sample injection of fluorescent analytes according to a Hadamard code sequence was performed using an optically gated sample injection technique, in which a sample plug was produced based on photodegradation by irradiation with an intense laser beam. The capillary and reservoirs were filled with a sample solution containing buffer components and SDS as a pseudostationary phase. A preliminary study confirmed that fluorescein ion could be photobleached in the presence of SIDS. The optically gated sample injection technique was then applied to multiple sample injection, based on a Hadamard matrix. The S/N in the electropherogram obtained by HT-MEKC was improved substantially compared to that obtained by a single injection method. When the technique was applied to the separation of several amino acids labeled with FITC, the S/N ratio for each amino acid was enhanced, without any evidence of degradation in separation resolution. Moreover, HT-MEKC was applied to the analysis of amino acids contained in a Japanese beverage, resulting in improved S/Ns for the amino acids.

DOI： 10.1002/elps.200600183

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