研究者詳細 - 金田　隆

論文 - 金田　隆

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A Sheath-Flow Electrochemical Detector for Capillary Electrophoresis 査読

Junji Inoue, Takashi Kaneta, Totaro Imasaka

Journal of Capillary Electrophoresis and Microchip Technology 10 ( 3&4 ) 69 - 73 2007年

On-column capture of a specific protein in capillary electrophoresis using magnetic beads 査読

Takashi Kaneta, Junji Inoue, Mototsugu Koizumi, Totaro Imasaka

ELECTROPHORESIS 27 ( 16 ) 3218 - 3223 2006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：WILEY

A method for capturing specific molecules separated by CE has been explored. To demonstrate on-column capture of migrating analyte molecules, two detection windows were fabricated on a capillary. Magnetic beads containing immobilized molecules that react with the specific molecules under study were placed between the detection windows in the capillary using magnets. Molecules in a sample solution injected into the capillary were separated and detected at the first detection window. After passing through the first detection window, the separated molecules encountered the magnetic beads, where the specific analyte was captured. As a result, the peak area for those analyte molecules decreased or disappeared completely at the second detection window. Rabbit IgG and carbonic anhydrase were employed to demonstrate on-column capture of a specific molecule. For rabbit IgG, magnetic beads containing the immobilized antibody (anti-rabbit IgG),were used. Rabbit IgG molecules were captured on the magnetic beads during CE migration. Furthermore, the capture of carbonic anhydrase was demonstrated by the reaction between magnetic beads (containing immobilized anti-rabbit IgG) and anti-carbonic anhydrase (rabbit IgG), before the beads were packed in the capillary. After packing the magnetic beads in the capillary, a mixture of two proteins was injected into the capillary. Two proteins were detected at the first detection window, while the peak corresponding to carbonic anhydrase disappeared at the second detection window. The results show that using an appropriate antibody, the present technique would be applicable to any proteins.

DOI： 10.1002/elps.200500936

Suppression of electroosmotic flow and its application to determination of electrophoretic mobilities in a poly(vinylpyrrolidone)-coated capillary 査読

T Kaneta, T Ueda, K Hata, T Imasaka

JOURNAL OF CHROMATOGRAPHY A 1106 ( 1-2 ) 52 - 55 2006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：ELSEVIER SCIENCE BV

A hydrophilic polymer, poly(vinylpyrrolidone) (PVP), was employed for suppressing the electroosmotic flow (EOF). A capillary was filled with aqueous PVP solution for coating the capillary wall with PVP; the PVP solution was then replaced by a migration buffer solution containing no PVP. Three types of PVP with different molecular weights were examined. The EOF was suppressed more effectively as the molecular weight of PVP increased. The EOF in the coated capillary was similar to 10-fold smaller than that of a bare capillary and was constant in the pH range of 6-8. The suppressed EOF was stable even when no PVP was added to the migration buffer. However, the EOF increased significantly when sodium dodecyl sulfate was added into the migration buffer. The method was applied for determining the electrophoretic mobilities of inorganic anions that have negative electrophoretic mobilities larger than the electroosmotic mobility of the bare capillary. A novel method for determining the electrophoretic mobilities was proposed based on the linear relationship between electric current and electrophoretic mobility. The electrophoretic mobility was proportional to the electric current. Therefore, the intercept of the regression equation represents the electrophoretic mobility at room temperature. The electrophoretic mobilities were in good agreement with the absolute electrophoretic mobilities. (c) 2005 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.chroma.2005.08.062

Hadamard transform capillary electrophoresis combined with laser-induced fluorometry using electrokinetic injection 査読

K Hata, T Kaneta, T Imasaka

ANALYTICA CHIMICA ACTA 556 ( 1 ) 178 - 182 2006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

Hadamard transform capillary electrophoresis (HTCE) based on electrokinetic injection allows laser-induced fluorescence detection using a small laser, namely the laser-diode-pumped YAG laser, as an excitation source. A small hole is fabricated at the center of a capillary by laser ablation; this hole functions as an inlet port for a sample solution. Therefore, the sample solution can be introduced electrophoretically into the capillary through the small hole. Multiple sample injection is accomplished by introducing a buffer solution from the end of the capillary and the sample solution through the hole. Both solutions are injected using two sets of high-voltage power supplies and migrate toward the opposite end of the capillary. A fluorescent analyte, rhodamine B, is successfully detected in the case of both single and multiple injection according to the Hadamard sequence code. By transforming the data encoded by the Hadamard matrix, the decoded data showed an increase in the signal-to-noise (S/N) ratio by a factor of 9.8. In the case of the sample containing two amino acids labeled with rhodamine B isothiocyanate (RBITC), although the concentration of every component including free RBITC is lower than the concentration limit of detection obtained by single injection, a substantial improvement in the sensitivity is achieved and all components are identified by the Hadamard transform technique. (c) 2005 Elsevier B.V. All fights reserved.

DOI： 10.1016/j.aca.2005.06.068

Capillary electrophoresis immunoassay based on an on-column immunological reaction 査読

S Miki, T Kaneta, T Imasaka

JOURNAL OF CHROMATOGRAPHY A 1066 ( 1-2 ) 197 - 203 2005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：ELSEVIER SCIENCE BV

An on-column immunological reaction was employed to achieve simple and rapid analysis in an immunoassay based on capillary electrophoresis using semiconductor laser-induced fluorescence detection. Human serum albumin (HSA) labeled with sulfoindocyanine succinimidyl ester (Cy5), a fluorescent compound with an absorption maximum at 649 urn, was used as a fluorescent probe for the immunoassay. In a binding assay, with anti-HSA as the analyte molecule, Cy5-HSA was injected in a capillary column followed by the injection of anti-HSA so as to form individual zones. By applying a potential, the anti-HSA reacted with Cy5-HSA at the boundary between Cy5-HSA and anti-HSA zones, since anti-HSA has a higher electrophoretic mobility than Cy5-HSA. Furthermore, the on-column method enhances the sensitivity by injecting a large volume of the sample. Free Cy5-HSA and its immunocomplex with anti-HSA were separated with less degradation in resolution than that predicted from the injection time of anti-HSA, even when the injection time for anti-HSA was increased. The ratio of the peak area of the complex to that of the total Cy5-HSA (free Cy5-HSA and the complex) increased in proportion to the injection time of anti-HSA. As a result, the detection limit was improved up to eight-fold (the concentration detection limit, 0.007 mg mL(-1)), for an injection time of 240 s, compared to that obtained using an off-column sample preparation. Furthermore, the on-column reaction method was applicable to an immunoassay to determine native HSA, in which native HSA and Cy5-HSA react with anti-HSA stepwise. The detection limit in the stepwise reaction immunoassay was 0.005 mg mL(-1), which is 14 times lower than that in an off-column method, with the analysis time less than 10 min as the result of increasing the injection time of native HSA. In addition, the present on-column immunoassay was applied to the sample containing a high concentration of salts for investigating the effect of salts in the sample solution. (C) 2005 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.chroma.2005.01.037

On-line concentration of a protein using denaturation by sodium dodecyl sulfate 査読

P Jing, T Kaneta, T Imasaka

ANALYTICAL SCIENCES 21 ( 1 ) 37 - 42 2005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：JAPAN SOC ANALYTICAL CHEMISTRY

A novel method for the on-column sample stacking of proteins is described. The strategy takes advantage of interactions between protein molecules and sodium dodecyl sulfate (SDS) monomers. A long plug of a protein sample (either acidic or basic) is injected into a capillary filled with a background electrolyte (BGE) containing SDS. When a potential is applied, the proteins interact with SDS monomers in the BGE to form protein-SDS complexes that migrate more slowly than the corresponding uncomplexed protein, resulting in protein stacking. Both acidic and basic proteins migrate at an almost identical electrophoretic velocity after stacking, which indicates that the protein-SDS complexes formed in the BGE zone have a similar charge/mass ratio. The mechanism of stacking was investigated using a sample consisting of a basic protein, lysozyme, and a small molecule, methylene blue. The findings clearly show that two interactions with SDS occur, a stepwise binding interaction between protein molecules and SDS monomers and an interaction in which the small molecules enter into micelles formed by SDS molecules. The method was also applied to the detection of a protein labeled with a fluorescent labeling reagent at trace levels. The labeled protein was detected even under labeling conditions where the labeling efficiency was too low to detect by short-plug injection.

DOI： 10.2116/analsci.21.37

On-line sample concentration techniques in capillary electrophoresis: Velocity gradient techniques and sample concentration techniques for biomolecules 査読

CH Lin, T Kaneta

ELECTROPHORESIS 25 ( 23-24 ) 4058 - 4073 2004年12月

記述言語：英語出版者・発行元：WILEY-V C H VERLAG GMBH

Methods with a high sensitivity and high separation efficiency are goals in analytical separation techniques. On-line sample concentration techniques in capillary electrophoresis (CE) separations have rapidly grown in popularity over the past few years because they achieve this goal. This review describes the methodology and theory associated with a number of different techniques, including electrokinetic and chromatographic methods. For small molecules, several on-line concentration methods based on velocity gradient techniques are described, in which the electrophoretic velocities of the analyte molecules are manipulated by field amplification, sweeping, and isotachophoretic migration, resulting in the on-line concentration of the analyte zones. In addition, the on-line concentration methods for macromolecules are described, since the techniques used for macromolecules (DNAs and proteins), are different from those for small molecules, with respect to either mechanism or methodology. Recent studies relating to this topic are also discussed, including electrophoretic and chromatographic techniques on capillary or microchip.

DOI： 10.1002/elps.200406172

Hadamard transform capillary electrophoresis combined with absorption spectrometry 査読

K Hata, T Kaneta, T Imasaka

ANALYTICAL CHEMISTRY 76 ( 15 ) 4421 - 4425 2004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

A novel injection device for applying absorption spectrometty to Hadamard transform (HT) capillary electrophoresis is described. A small hole, at the center part of the capillary, functions as an inlet port for the sample. The hole is immersed in a sample solution and the end of the capillary that is usually employed for sample introduction is immersed in a buffer solution. An ultraviolet absorption detector is placed between the sample injection port and the other end of the capillary filled with a buffer solution. A high potential is continuously applied between the injection port and the end of the capillary, which allows the sample solution to be introduced into the separation capillary. By application of a higher potential modulated according to a Hadamard code between both ends of the capillary, the buffer solution is injected into the separation capillary. In some preliminary experiments, this injection device was utilized to introduce a single sample segment into a capillary. As expected, a single peak was observed in the electropherogram for a sample containing a single component. This device was then employed for multiple sample injection in HT capillary electrophoresis. An 8-fold improvement in the S/N ratio was observed when the HT technique was used, in which a 255-order of a Hadamard matrix was used, as expected from theory. The present approach was also utilized for the sensitive detection of a sample comprised of multiple components.

DOI： 10.1021/ac035366j

Detection of 27 molecules in a single injection volume by hadamard transform capillary electrophoresis 査読

T Kaneta, K Kosai, T Imasaka

ANALYTICAL SCIENCES 19 ( 12 ) 1659 - 1661 2003年12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：JAPAN SOC ANALYTICAL CHEMISTRY

A concentration detection limit of 100 fM was achieved for the fluorescein ion by improving the experimental setup used for Hadamard transform capillary electrophoresis. Two argon-ion lasers, a gating laser for sample injection and a probe laser for the excitation of analyte molecules, were employed for the efficient photodegradation of analyte molecules in laser-induced fluorescence detection using an optically gated sample-injection method. In addition, a dichroic mirror, located in the pathway of the probe laser was used to exclude the other lines of the argon-ion laser. Using a Hadamard matrix on the order of 2046, the concentration limit of detection for fluorescein ion was determined to be 100 fM at S/N = 3, in which the average number of molecules in a single injection volume was calculated to be 27. The influences of the output power in both the gating and probe lasers on the sensitivity are also discussed.

DOI： 10.2116/analsci.19.1659

Application of Hadamard transformation to chiral separation by capillary electrophoresis 査読

T Kaneta, M Nishida, T Imasaka

BUNSEKI KAGAKU 52 ( 12 ) 1193 - 1197 2003年12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：JAPAN SOC ANALYTICAL CHEMISTRY

Hadamard transform capillary electrophoresis, which is a sensitive detection method using multiple sample injection, was applied to the separation of glutamic acid enantiomers labeled with fluorescein isothiocyanate (FITC). A capillary was filled with a sample solution containing FITC-labeled D,L-glutamic acids, cyclodextrin, and buffer components. The sample was injected by an optically gated sample injection method, in which an intense laser beam was focused onto the capillary to convert fluorescent analyte ions to nonfluorescent ones by photodegradation. To form analyte plug, a shutter placed in the pathway of the laser beam was closed for a constant time period. In the presence of cyclodextrin, enatiomers of FITC-labeled glutamic acid could be injected by the optically gated sample injection method. Two cyclodextrins, beta- and gamma-cyclodextrins, were employed as chiral selectors in the capillary electrophoretic separations. For the separation of FITC-labeled glutamic acid enantiomers, gamma-cyclodextrin shows a better resolution than P-cyclodextrin. The optimum concentration of gamma-cyclodextrin was found to be 10 mM. Multiple sample injection was accomplished by closing or opening the shutter according to a sequence based on a Hadamard matrix. When the resulting electropherogram was transformed by the inverse Hadamard matrix, The S/N ratio in the transformed result was enhanced with no degradation in the resolution, compared to that in the electropherogram obtained by a single injection method. As a result, Hadamard transform capillary electrophoresis allows a significant improvement in the SIN ratios for FITC-labeled glutamic acid enantionters. Therefore, the present method will be applicable to the separation and sensitive detection of enantiomers labeled with FITC.

DOI： 10.2116/bunsekikagaku.52.1193

Absorption spectrometry using a diffractive optical element constructed by gel electrophoresis 査読

A Shigyo, S Miki, T Kaneta, T Imasaka

ANALYTICAL SCIENCES 19 ( 5 ) 795 - 798 2003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：JAPAN SOC ANALYTICAL CHEMISTRY

A novel diffractive optical element was constructed by means of gel electrophoresis. Five capillaries were filled with a methylene blue solution and inserted into a slab gel. An electrical potential was then applied to the slab gel. Methylene Blue in the capillaries underwent electrophoretic migration, forming clear blue bands in the gel. The bands functioned as a transmission grating, and diffracted a laser beam. The intensity of the diffracted light increased with increasing the concentration of methylene blue.

DOI： 10.2116/analsci.19.795

Hadamard transform microchip electrophoresis combined with diode laser fluorometry 査読

K Hata, Y Kichise, T Kaneta, T Imasaka

ANALYTICAL CHEMISTRY 75 ( 7 ) 1765 - 1768 2003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：AMER CHEMICAL SOC

The application of a Hadamard transform technique to microchip electrophoresis is described. The sample is electrokinetically injected into a separation channel and is then detected by diode laser-induced fluorometry. The sample and buffer solutions are introduced into the channel by controlling the high voltages applied to the solutions, according to a code determined by a Hadamard matrix. The S/N ratio of the signal in the electropherogram can be improved by a factor of 5 in comparison with that obtained by a conventional single-injection method, although an 8-fold improvement is theoretically predicted when a 255-order matrix is used.

DOI： 10.1021/ac026330e

Determination of dye/protein ratios in a labeling reaction between a cyanine dye and bovine serum albumin by micellar electrokinetic chromatography using a diode laser-induced fluorescence detection 査読

P Jing, T Kaneta, T Imasaka

ELECTROPHORESIS 23 ( 15 ) 2465 - 2470 2002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：WILEY-V C H VERLAG GMBH

The degree of labeling, i.e., dye/protein ratio (DIP) is important for characterizing properties of dye labeling with proteins. A method for the determination of this ratio between a fluorescent cyanine dye and bovine serum albumin (BSA), based on the separation of the labeling mixture using micellar electrokinetic chromatography with diode laser-induced fluorescence detection, is described. Two methods for the determination of DIP were examined in this study. In these methods, a hydrolysis product and impurities, which are usually unfavorable compounds that are best excluded for protein analysis, were utilized to determine the amounts of dye bound to BSA. One is a direct method in which a ratio of the peak area of BSA to the total peak area of all the products produced in the labeling reaction was used for determining the average number of dye molecules bound to a single BSA molecule. The other is an indirect determination, which is based on diminution of all peak areas related to the products except for the labeled BSA. These methods were directly compared by means of a spectrophotometric method. The experimental results show that the indirect method is both reliable and sensitive. Therefore, DIP values can be determined at trace levels using the indirect method.

DOI： 10.1002/1522-2683(200208)23:15<2465::AID-ELPS2465>3.0.CO;2-G

Band broadening caused by the multiple labeling of proteins in micellar electrokinetic chromatography with diode laser-induced fluorescence detection 査読

P Jing, T Kaneta, T Imasaka

JOURNAL OF CHROMATOGRAPHY A 959 ( 1-2 ) 281 - 287 2002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 SCIENCE BV

When a labeling reagent is used, in the determination of proteins by capillary electrophoresis with laser-induced fluorescence detection, the multiple labeling of proteins frequently occurs, which can degrade the separation efficiency. In order to understand the influence of the multiple labeling of proteins on separation efficiency, the band broadening caused by a labeling reaction between bovine serum albumin (BSA) and a cyanine fluorescent dye (Cy5) was investigated using micellar electrokinetic chromatography in conjunction with diode laser-induced fluorometry. With the aid of an internal standard, methylene blue, the height equivalent to the theoretical plate (HETP) ratio of BSA to methylene blue was used as an indicator for band broadening under optimum separation conditions. Labeling conditions, including reaction buffer pH, reaction time, and initial concentration of Cy5 to bovine serum albumin, were found to influence the HETP ratio. The separation efficiency for the labeled protein was degraded by experimental conditions employed in the labeling, which indicates an increase in the heterogeneity of the final products. (C) 2002 Elsevier Science B.V. All rights reserved.

DOI： 10.1016/S0021-9673(02)00357-6

Ultratrace analysis based on Hadamard transform capillary electrophoresis 査読

T Kaneta, K Kosai, T Imasaka

ANALYTICAL CHEMISTRY 74 ( 10 ) 2257 - 2260 2002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 CHEMICAL SOC

Hadamard transform capillary electrophoresis, which is based on a multiple sample injection technique, was combined with laser-induced fluorometry and utilized in the determination of analytes at subpicomolar levels. The sensitivity was substantially improved by increasing the order, i.e., the number of elements, of the Hadamard matrix. In facts the signal-to-noise ratio was enhanced 18-fold by the use of a matrix of order 2047. A feasibility study was carried out by computer simulation to study the detection of an average of less than a single molecule in a single injection volume. The signal peak was clearly observable even under conditions at which only 0.3 molecule is present in the volume. Thus, this approach is potentially useful for ultratrace analysis, in which conventional single-injection capillary electrophoresis cannot be applied.

DOI： 10.1021/ac011149b

Micellar electrokinetic chromatography with diode laser-induced fluorescence detection as a tool for investigating the fluorescence labeling of proteins 査読

P Jing, T Kaneta, T Imasaka

ELECTROPHORESIS 23 ( 4 ) 550 - 555 2002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

We describe the use of micellar electrokinetic chromatography with diode laser-induced fluorometry (MEKC-DLIF) as a tool to characterize a labeling reaction between sulfoindocyanine succinimidyl ester (Cy5), a cyanine fluorescent dye, and a model protein, bovine serum albumin (BSA). To decrease the influence of imprecise injection, methylene blue was added as an internal standard. Labeled BSA was completely separated from the unconjugated Cy5, and methylene blue under optimized conditions. A kinetic study of the reaction was performed by changing some parameters, such as reaction buffer pH, reaction buffer concentration and the initial concentration of BSA. A comparison between the current method and traditional methods was made.

DOI： 10.1002/1522-2683(200202)23:4<550::AID-ELPS550>3.0.CO;2-F

An "optical channel": A technique for the evaluation of biological cell elasticity 査読

T Kaneta, J Makihara, T Imasaka

ANALYTICAL CHEMISTRY 73 ( 24 ) 5791 - 5795 2001年12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

The development of an optical channel, a new analytical technique for evaluating the elasticity of biological cells, is described in this study. Two types of erythrocyte cells, i.e., young and old cells, were examined via their introduction into a flowing medium, to which a laser beam was focused in the opposite direction. An erythrocyte cell is trapped in a laser beam by a gradient force, moves in the downstream direction, and is then elongated at the beam waist. The change in shape was measured directly using a microscope equipped with a charge-coupled-device camera. It is probable the main driving force for the cell deformation is a shear stress generated by a medium flow, since an estimate of the gradient force suggests that it is too small to change the shape of an erythrocyte. The average values of the elongation of young and old cells were 2.4 +/- 0.6 and 2.1 +/- 0.5, respectively. These values are in reasonably good agreement with values obtained using a rheoscope method. The deformation was measured without any physical contact to the solid surface, and therefore, damage to cells such as these are minimal.

DOI： 10.1021/ac010441g

Hadamard transform capillary electrophoresis. 査読

T Kaneta

ANALYTICAL CHEMISTRY 73 ( 19 ) 540A - 547A 2001年10月

Immunoassay for human serum albumin using capillary electrophoresis-semiconductor laser-induced fluorometry 査読

S Miki, T Kaneta, T Imasaka

JOURNAL OF CHROMATOGRAPHY B 759 ( 2 ) 337 - 342 2001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

Capillary electrophoresis combined with semiconductor laser-induced fluorometry was applied to an immunoassay of human serum albumin. Human serum albumin was labeled with a fluorescent molecule (Cy5), which has an absorption maximum at 649 nm. The labeled albumin was purified by ultrafiltration in order to reduce signals, which are unreacted labeling reagent, product, and fragment products derived therefrom. After the purification, no signal for unreacted labeling reagent and fragment products was detectable in the electropherogram of the labeled albumin. The labeled albumin was then reacted with anti-albumin to form an immunocomplex, which was separated from the excess free albumin. The competitive immunoassay was used in the determination of human serum albumin in a controlled serum sample, using die labeled albumin. The obtained value was found to be 0.21 +/-0.02 mg/ml, which is in good agreement with other known values. (C) 2001 Elsevier Science B.V. All rights reserved.

DOI： 10.1016/S0378-4347(01)00228-6

Migration Behavior of Alkylphenols, Bisphenol A and Bisphenol S Studied by Capillary Electrophoresis Using Sulfated β-Cyclodextrin 査読

Analytical Sciences 17 ( 6 ) 763 - 768 2001年6月

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