Papers - Fujimoto, Keiji

DNA-carring latex particles for DNA diagnosis 3. Detection of point mutation DNA using an indirect labeled probe

Colloids and Surfaces A: Physicochemical and Engineering Aspects (Elsevier) vol.153   445-451 1999

Joint Work, Accepted

Dispersion polymerization of methyl methacrylate using macro-azo-initiator

Nakamura K., Fujimoto K., Kawaguchi H.

Colloids and Surfaces A: Physicochemical and Engineering Aspects (Elsevier) (Colloids and Surfaces A: Physicochemical and Engineering Aspects)  vol.153 ( 1-3 ) 195-201 - 201 1999

Research paper (scientific journal), Joint Work, Accepted,  ISSN  09277757

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Poly(dimethylsiloxane)-containing macro-azo-initiator was used in dispersion polymerization of methyl methacrylate. Stable microspheres were obtained when the solvents had solubility parameters in the range of 7.4-8.2. The microspheres were stabilized by a stabilizer formed in-situ, that is, polydimethylsiloxane-block-poly(methyl methacrylate). Copyright (C) 1999 Elsevier Science B.V. All rights reserved.

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Preparation of unsymmetrical microspheres at the interfaces

Fujimoto K., Nakahama K., Shidara M., Kawaguchi H.

Langmuir (Americal Chemical Society) (Langmuir)  vol.15 ( 13 ) 4630-4635 - 4635 1999

Research paper (scientific journal), Joint Work, Accepted,  ISSN  07437463

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Many synthetic microspheres are spherical and symmetrical because of the thermodynamical limitations of the reaction systems. We developed modification techniques to prepare spherical microspheres with intrinsic unsymmetry, for example, a microsphere with both an anionic and a cationic part. The modification was performed at the liquid-solid interface. Reactive microspheres with p-nitrophenyl moieties were settled onto the IgG-preadsorbed substrate, and the reaction between the activated ester moieties and IgG molecules proceeded only at the interface upon attachment. After the reaction, the modified microspheres were detached from the substrate with ultrasonication. In another modification at the air-water interface, the dispersion of reactive microspheres in ethanol was first spread on a water surface to produce the monolayer of microspheres, and then hydrolysis at the water side of the monolayer was performed by adding a NaOH aqueous solution to the subphase. After this unsymmetrical hydrolysis, we carried out IgG immobilization with the remaining reactive moieties. In both interface methods, the immobilized IgG molecules were labeled with colloidal gold particles conjugated with anti-IgG antibodies. Transmission electron microscopy indicated that gold particles could be observed on only the immobilized side. Small microspheres were coupled to the reactive microsphere, and its unsymmetry was confirmed by scanning electron microscopy. Such unsymmetrical microspheres will be applicable to the electrical rheology, diagnosis, display technology, and creation of functional devices through the assembling of microspheres.

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Smart latex es for bioseparation

Kawaguchi H., Fujimoto K.

Bioseparation (Kluwer Academic Publishers) (Bioseparation)  vol.7 ( 4-5 ) 253-258 - 258 1999

Research paper (scientific journal), Joint Work, Accepted,  ISSN  0923179X

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Monodisperse, thermosensitive microspheres with sub-micron diameters are used for separation and collection of proteins and other biomolecules. The thermosensitivity gives the microspheres two valuable features. One is the controllability of affinity between microsphere and protein with temperature. The quality and quantity of proteins to be adsorbed on the microspheres can be controlled with temperature. The other feature is reversible control of dispersion stability. Microspheres which have adsorbed target proteins in the dispersion can be easily recovered by changing temperature.

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Stimuli-Response of Microsphere Having Poly(N-isopropylacrylamide) Shell

Matsuoka H., Fujimoto K., Kawaguchi H.

Polymer Journal (The Society of Polymer Science, Japan) (Polymer Journal)  vol.31 ( 11 pt 2 ) 1139-1144 - 1144 1999

Research paper (scientific journal), Joint Work, Accepted,  ISSN  00323896

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Several kinds of submicron-sized microspheres having environment-sensitive shells were prepared and the effect of shell structure on the properties of microspheres was studied. The shell was composed of crosslinked or hairy poly(N-isopropyl acrylamide) (PNIPAM) whose lower critical solution temperature was about 32 °C but depended on the architecture of the shell. The resposivity of the shell to the change in environmental condition was sluggish when the shell was crosslinked. The decrease in the crosslink density of the shell was slightly effective in sharpening the response. But even non-crosslinked, hairy PNIPAM shell did not exhibit discontinuous transition at the critical condition. The response of hairy PNIPAM shell to environmental change was drastically sharpened by incorporating a slight amount of acrylic acid (AAc) in the hair.

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異方性微粒子の作製とそのはたらき

藤本啓二、中濱数理、設楽美和子、川口春馬

ポバール会記録 vol.110   19-25 1998.12

Joint Work

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微粒子に新たな自由度を与えるための一つの方策として、微粒子への異方性の導入を提案した。このような異方性処理によって上半球がプラス、下半球がマイナスといったものや、ある部分にだけ特定の高分子鎖が生えたものといった両親媒性を有するものが作製できると考えられる。この両親媒性は一個の微粒子に運動性をはじめとする様々な特異性を付与するために有効であろう。

アニオン性ポリマーによるマクロファージの活性化

藤本啓二、向井章、川口春馬、竹内勤

高分子加工 vol.47 ( No.8 ) 11-15 1998.08

Research paper (scientific journal), Joint Work

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合成したアニオン性ポリマーについて、その免疫系（主にマクロファージ）への作用の一端を知ることができた。特に寄生虫感染の初期においてはこのポリマーはマクロファージ等の活性化を誘導して防御に努めている。しかしその効果は長くは続かず、その後は主に細胞性免疫系の活性化を通して本質的な抗寄生虫活性を発現できるようになると考えられた。

DNA-carrying latex particles for DNA diagnosis 2. Distinction of normal and point mutant DNA using S1 nuclease

Hatakeyama M., Nakamura K., Iwato S., Handa H., Fujimoto K., Kawaguchi H.

Colloids and Surfaces B: Biointerfaces (Colloids and Surfaces B: Biointerfaces)  10 ( 3 ) 171 - 178 1998.02

Research paper (scientific journal), Accepted,  ISSN  09277765

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In our previous paper, we reported a DNA diagnosis system in which fully complementary DNA was selectively hybridized with a DNA immobilized on latex particles but one-point mutant DNA was not. The system, however, included a few disadvantages. To overcome them, a single-strand DNA cleaving enzyme (S1 nuclease) was used in this study. S1 nuclease cleaved the unhybridized and loosely hybridized DNA's on the particles and enabled the determination of the fully complementary hybrid. This method saved us a troublesome labeling of the samples.

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Monodisperse Microspheres Exhibiting Discontinuous Response to Temperature Change

Matsuoka,H., Fujimoto,K., Kawaguchi,H.

Polymer Gel Network, 6(1998) (Polymer Gels and Networks)  6 ( 5 ) 319-332 - 332 1998

Research paper (scientific journal), Joint Work, Accepted,  ISSN  09667822

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これまで作製されてきた感温性ゲル微粒子のサイズ変化は連続的であった。そこで架橋点をなくするためにコア微粒子からのN-イソプロピルアクリルアミドのグラフト重合を行った。さらにモノマーにアクリル酸を少量加えることにより不連続なサイズ変化を示す微粒子の作製が出来るようになった。

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DNA-carrying latex particles for DNA diagnosis. 1. Separation of normal and point mutant DNAs according to the difference in hybridization efficiency

Hatakeyama,M., Iwato,S., Fujimoto,K., Handa,H., and Kawaguchi,H.

Colloids and Surfaces B: Biointerfaces, (Colloids and Surfaces B: Biointerfaces)  vol.10 ( 3 ) 161-169 - 169 1998

Research paper (scientific journal), Joint Work, Accepted,  ISSN  09277765

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正常なDNA鎖同士のハイブリダイゼーションと片方のDNAにだけ点変異がある場合のハイブリダイゼーションとでは、後者のほうが結合力が弱いことがわかった。そこで微粒子にDNA一本鎖を固定化して、点変異DNAだけが検出できるように種々の条件の設定を行い、その検出に成功した。

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DNA-carrying latex particles for DNA diagnosis. 2. Distinction of normal and point mutant DNAs using S1 nuclease

Hatakeyama,M., Nakamura,K., Iwato,S., Fujimoto,K., Handa,H., and Kawaguchi,H.

Colloids and Surfaces B: Biointerfaces vol.10   171-178 1998

Joint Work, Accepted

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点変異があるDNAとその変異点以外は相補的なDNAとの間では不完全ではあるが二本鎖が形成される。この不完全部分を認識して切断する酵素を用いて、点変異をもつDNAの検出に成功した。

Activation of neutrophil-like cells stimulated by polymer surfaces

Fujimoto,K., Sasaki,J., Tomida,Y., Wakabayashi,Y., and Kawaguchi,H.

高分子論文集 (Kobunshi Ronbunshu)  vol.55 ( No.6 ) 367-371 - 371 1998

Research paper (conference, symposium, etc.), Joint Work, Accepted,  ISSN  03862186

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様々な高分子表面による好中球の活性化について検討を行った。電荷を導入した表面および疎水性の表面は一過性であるが強く細胞を活性化させ、親水性の表面は弱いが持続的に細胞を活性化させることがわかった。

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Inverse microemulsion polymerization of acrylamide in the presence of N,N-dimethylacrylamide

Juranicova,V., Kawamoto,S., Fujimoto,K., Kawaguchi,H., and Barton,J.

Die Angewandte Makromolekulare Chemie (Angewandte Makromolekulare Chemie)  vol.258   27-31 - 31 1998

Research paper (scientific journal), Joint Work, Accepted,  ISSN  00033146

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N,N-ジメチルアクリルアミド存在下でアクリルアミドの逆相マイクロエマルジョン重合を行った。

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高分子表面によるタンパク質のリフォールディング制御

藤本啓二、清水秀信、川口春馬

化学と工業 vol.50 ( No.10 ) 1523-1525 1997.10

Research paper (scientific journal), Joint Work

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生体内でつくられたタンパク質の鎖は、折り畳まれて活性を示す立体構造をとるようになる。最近、この折り畳みを助ける生体システムについて研究が進んでいる。我々は初めてこの折り畳みの人工システムを考案・作製し、実際にタンパク質を効率よくフォールディングさせることに成功した。

人工材料表面によるタンパク質のリフォールディング制御

藤本啓二、清水秀信、川口春馬

ポバール会記録 vol.110   19-25 1997.07

Joint Work

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生体内でつくられたタンパク質の鎖は、折り畳まれて活性を示す立体構造をとるようになる。最近、この折り畳み（リフォールディング）を助ける生体システムについて研究が進んでいる。我々はこの折り畳みの人工システムを考案・作製し、タンパク質のリフォールディング反応を促進させることに成功した。

Cell activation by the micropatterned surface with settling particles

Fujimoto K., Takahashi T., Miyaki M., Kawaguchi H.

Journal of Biomaterials Science, Polymer Edition (Journal of Biomaterials Science, Polymer Edition)  8 ( 11 ) 879 - 891 1997.01

Research paper (scientific journal), Accepted,  ISSN  09205063

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Surface topography plays an important role in cell orientation and morphogenesis. In this study, we prepared a micropatterned surface with settling particles to obtain more detailed information about the cell recognition against the microstructured surface. Core-shell type particles having a poly-(N-isopropylacrylamide) (polyNIPAM) shell were prepared by seeded polymerization. Particles were settled on a polystyrene (PSt) flat dish by the spinner to prepare a micropatterned surface with settling particles. It could be seen that the polyNIPAM shell shrunk above and swelled below the LCST. For comparison, a thermosensitive flat surface was prepared by the graft polymerization of NIPAM. No morphologic change of cells contacting the both surfaces was observed with either an optical or a scanning electron microscope. Moreover, particles could move or roll on these surfaces when shaking the dishes. The weak interaction between neutrophil-like cells and the micropatterned surface with settling particles or the polyNIPAM-grafted surface was estimated by measurement of active oxygen released by cells. A little release could be observed at both 25 and 35°C. The amount of released active oxygen at 35°C was slightly larger than at 25°C. When the temperature was suddenly changed, the dynamic changes of particle shape and size resulted in the excess release of active oxygen from cells contacting the micropatterned surface with settling particles. Meanwhile no stimulation could be observed in the polyNIPAM-grafted surface even if the temperature is suddenly changed. These results indicate that the micropatterned surface with settling particles can induce the dynamic stimulus at a patterned input mode. © VSP 1997.

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Preparation of dimerized polypeptide-carrying microspheres and purification of specific proteins bound to these microspheres.

Hatakeyama,M., Ogura,Y., Sawada,J., Fujimoto,K., Handa,H., and Kawaguchi,H.

Colloids and Surfaces B: Biointerfaces (Colloids and Surfaces B: Biointerfaces)  vol.10 ( 1 ) 41-49 - 49 1997

Research paper (scientific journal), Joint Work, Accepted,  ISSN  09277765

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二量化したペプチド断片の微粒子への固定化を行った。この微粒子を用いて二量化ペプチド断片と結合する核内タンパク質の単離を試みた。

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Activity of enzymes immobilized on microspheres with thermosensitive hairs

Yasui,M., Shiroya,T., Fujimoto,K., and Kawaguchi,H.

Colloids and Surfaces B: Biointerfaces (Colloids and Surfaces B: Biointerfaces)  vol.8 ( 6 ) 311-319 - 319 1997

Research paper (scientific journal), Joint Work, Accepted,  ISSN  09277765

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微粒子表面の感温性のポリマー鎖に酵素の固定化を行った。この微粒子は温度変化により３つのサイズに変化した。それぞれの状態で酵素活性は異なり、温度による制御の可能性が示唆された。

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Ex vivo and in vivo evaluation of the blood compatibility of surface-modified polyurethane catheters

Inoue,H., Fujimoto,K., Uyama,Y., and Ikada,Y.

Journal of Biomedical Materials Research (Journal of Biomedical Materials Research)  vol.35 ( 2 ) 255-264 - 264 1997

Research paper (scientific journal), Joint Work, Accepted,  ISSN  00219304

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ポリエステルウレタンをオゾン処理し、水溶性でノニオン性のポリマーのグラフト化を行った。この改質ポリウレタンの血液適合性を調べたところ、良好な結果を得ることが出来た。

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Affinity Latex for DNA Diagnosis

Kawaguchi,H., Hatakeyama,M., Iwato,S., Fujimoto,K., Handa,H.

Advances in Polymeric Biomaterials Science,Akaike,T., Okano,T., Akashi,M., Terano,M., and Yui,N., eds.CMC Inc., Tokyo    355-362 1997

Joint Work, Accepted

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微粒子にDNAを固定化することにより遺伝子診断のデバイスとしての応用を試みた。