研究者紹介システム

山地 秀樹
ヤマジ ヒデキ
大学院工学研究科 応用化学専攻
教授
応用化学関係
Last Updated :2022/06/23

研究者情報

所属

  • 【主配置】

    大学院工学研究科 応用化学専攻
  • 【配置】

    工学部 応用化学科, 先端バイオ工学研究センター

学位

  • 博士(工学), 京都大学
  • 昆虫細胞を用いた有用物質生産

授業科目

ジャンル

  • 科学・技術 / 工学

コメントテーマ

  • バイオリアクター
  • 昆虫細胞培養による有用物質生産
  • バイオ燃料
  • バイオ医薬品

研究活動

研究分野

  • ものづくり技術(機械・電気電子・化学工学) / バイオ機能応用、バイオプロセス工学

受賞

  • 2018年11月 Japanese Association for Animal Cell Technology (JAACT), Best Poster Presentation Award, The 31st Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2018 Tsukuba), Production of influenza virus-like particles in stably transformed insect cells

    MATSUDA Takuya, TANIJIMA Toshikazu, Kyoko MASUMI-KOIZUMI, KATSUDA Tomohisa, YAMAJI Hideki

    国際学会・会議・シンポジウム等の賞

  • 2018年11月 Japanese Association for Animal Cell Technology (JAACT), Best Poster Presentation Award, The 31st Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2018 Tsukuba), Production of antibody Fab fragment using 2A peptide in insect cells

    Yu Mizote, Kyoko Masumi-Koizumi, Tomohisa Katsuda, Hideki Yamaji

  • 2017年09月 化学工学会, バイオ部会優秀ポスター賞, 昆虫細胞による2Aペプチドを用いた抗体タンパク質の生産

    溝手 結, 増見 恭子, 勝田 知尚, 山地 秀樹

    国内学会・会議・シンポジウム等の賞

  • 2016年11月 Japanese Association for Animal Cell Technology (JAACT), Best Poster Presentation Award, Production of virus-like particles using recombinant insect cells

    TANISHIMA Toshikazu, KATSUDA Tomohisa, YAMAJI Hideki

    国際学会・会議・シンポジウム等の賞

  • 2010年10月 Asia Pacific Confederation of Chemical Engineering, Best Poster Paper Award (The 13th Asia Pacific Confederation of Chemical Engineering Congress (APCChE 2010)), Production of Japanese encephalitis virus-like particles in insect cell expression systems

    YAMAJI Hideki, NAGASUGA Takashi, TAKAHASHI Yusuke, NAKAMURA Masataka, KATSUDA Tomohisa, KUWAHARA Miwa, KONISHI Eiji

    国際学会・会議・シンポジウム等の賞

  • 2010年02月 化学工学会関西支部, 化学工学会バイオ部会, 第8回最先端バイオテクノロジー発表会ポスター賞, 一本鎖抗体の可溶性発現に及ぼすSD配列改変の影響, 野上達弘

    野上 達弘, 木原 麻那, 佐野 宰久, 勝田 知尚, 山地 秀樹

  • 2009年12月 Young Asian Biochemical Engineers’ Community, Best Poster Presentation Award for an Original Contribution to the 15th Symposium of Young Asian Biochemical Engineers’ Community, Effects of Shine-Dalgarno sequence on the production of single-chain Fv antibody by Escherichia coli

    KIHARA Mana, NOGAMI Tatsuhiro, SANO Tadahisa, KATSUDA Tomohisa, YAMAJI Hideki

  • 1997年09月 化学工学会, 平成8年度化学工学会賞奨励賞 (内藤雅喜記念賞), 多孔性の保持粒子を用いる浮遊性動物細胞の固定化培養に関する研究

    山地 秀樹

論文

  • Ryou Nakanuma, Kyoko Masumi-Koizumi, Yuki Ohmuro-Matsuyama, Tomohisa Katsuda, Hideki Yamaji

    Springer Science and Business Media {LLC}, 2021年06月29日, Cytotechnology, 73 (3), 299 - 305

    [査読有り]

    研究論文(学術雑誌)

  • Takuya Matsuda, Toshikazu Tanijima, Kyoko Masumi-Koizumi, Tomohisa Katsuda, Hideki Yamaji

    Influenza vaccines have long been manufactured in embryonated chicken eggs. This method has some problems such as a long production period (about 6 months) and use of large amounts of infectious pathogens. Recently, the production of recombinant subunit vaccines using the baculovirus–insect cell system has been extensively investigated. In this system, viral immunodominant components can be produced more rapidly and in a larger scale than in the conventional egg-based process. However, continuous production is virtually impossible because infection of recombinant baculovirus results in the death of host insect cells. In the present study, we established stably transformed insect cells that secreted influenza virus-like particles (VLPs) consisting of hemagglutinin (HA), the major protective antigen of influenza A virus, and matrix protein 1 (M1), another structural protein of the virus. Hemagglutination assay and transmission electron microscopy (TEM) suggested that HA produced by recombinant insect cells kept the hemagglutination activity and the morphology of the VLPs was similar to that of wild type influenza virus particles.

    {EDP} Sciences, 2021年01月, MATEC Web of Conferences, 333, 07009 - 07009

    [査読有り]

    研究論文(学術雑誌)

  • Takuya Matsuda, Toshikazu Tanijima, Akito Hirose, Kyoko Masumi-Koizumi, Tomohisa Katsuda, Hideki Yamaji

    Elsevier {BV}, 2020年11月, Biochemical Engineering Journal, 163, 107757 - 107757, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Yu Mizote, Kyoko Masumi-Koizumi, Tomohisa Katsuda, Hideki Yamaji

    Elsevier {BV}, 2020年08月, Journal of Bioscience and Bioengineering, 130 (2), 205 - 211, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Production of influenza virus-like particles in insect cells

    Hideki Yamaji, Takuya Matsuda, Toshikazu Tanijima, Kyoko Masumi-Koizumi, Tomohisa Katsuda

    2020年05月, BMC Proceedings, 14, 14 - 15, 英語

    研究論文(国際会議プロシーディングス)

  • OHMURO-MATSUYAMA Yuki, KATSUDA Tomohisa, YAMAJI Hideki

    2019年01月, In Vitro Cellular & Developmental Biology - Animal, 55 (1), 1 - 6, 英語

    [査読有り]

    研究論文(学術雑誌)

  • OHMURO-MATSUYAMA Yuki, GOMI Keiko, YAMAJI Hideki, YAMASHITA Takahiro, UEDA Hiroshi

    2018年12月, Analytical Biochemistry, 563 (1), 61 - 66, 英語

    [査読有り]

    研究論文(学術雑誌)

  • YAMAJI Hideki, TANIJIMA Toshikazu, MATSUDA Takuya, Kyoko MASUMI-KOIZUMI, KATSUDA Tomohisa

    2018年10月, New Biotechnology, 44S, S159, 英語

    研究論文(国際会議プロシーディングス)

  • Yuki Ohmuro-Matsuyama, Hideki Yamaji

    Monoclonal antibodies and antibody fragments have recently been developed for use in diverse diagnostic and therapeutic applications. Insect cells can efficiently secrete recombinant proteins such as antibody molecules through post-translational processing and modifications that are similar to those performed in mammalian cells. In eukaryotic cells, the signal sequence in a nascent polypeptide is recognized by the signal recognition particle, and the polypeptide is then folded and modified in the endoplasmic reticulum. The signal sequence consists of three regions, a positively charged N-terminus, a hydrophobic core, and a polar C-terminus. In the present study, we examined the substitutions of the characteristic amino acids of a Drosophila immunoglobulin heavy chain binding protein signal sequence, and investigated the effect on the secretory production of an antibody Fab fragment from lepidopteran insect cells in transient expression. A modification of the signal sequence for the heavy chain resulted in a twofold increase in the secreted Fab fragment, while the modification for the light chain led to a more than 3.6-fold increase.

    Springer Netherlands, 2018年06月01日, Cytotechnology, 70 (3), 891 - 898, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Shun Tamaki, Mitsuhiko Yagi, Yuki Nishihata, Hideki Yamaji, Yasushi Shigeri, Tomohide Uno, Hiromasa Imaishi

    The aromatic compound p-hydroxybenzoate (PHBA) is an important material with multiple applications, including as a building block of liquid crystal polymers in chemical industries. The cytochrome P450 (CYP) enzymes are beneficial monooxygenases for the synthesis of chemicals, and CYP53A15 from fungus Cochliobolus lunatus is capable of executing the hydroxylation from benzoate to PHBA. Here, we constructed a system for the bioconversion of benzoate to PHBA in Escherichia coli cells coexpressing CYP53A15 and human NADPH-P450 oxidoreductase (CPR) genes as a redox partner. For suitable coexpression of CYP53A15 and CPR, we originally constructed five plasmids in which we replaced the N-terminal transmembrane region of CYP53A15 with a portion of the N-terminus of various mammalian P450s. PHBA productivity was the greatest when CYP53A15 expression was induced at 20°C in 2×YT medium in host E. coli strain ΔgcvR transformed with an N-terminal transmembrane region of rabbit CYP2C3. By optimizing each reaction condition (reaction temperature, substrate concentration, reaction time, and E. coli cell concentration), we achieved 90% wholecell conversion of benzoate. Our data demonstrate that the described novel E. coli bioconversion system is a more efficient tool for PHBA production from benzoate than the previously described yeast system.

    Korean Society for Microbiology and Biotechnology, 2018年03月01日, Journal of Microbiology and Biotechnology, 28 (3), 439 - 447, 英語

    [査読有り]

    研究論文(学術雑誌)

  • YAMAJI Hideki, MORI Keita, HAMADA Hirotsugu, OHMURO-MATSUYAMA Yuki, KATSUDA Tomohisa

    2018年03月, BMC Proceedings, 12 (3), 19, 英語

    研究論文(国際会議プロシーディングス)

  • Yuki Ohmuro-Matsuyama, Takahiro Yamashita, Huan Lin, Hideki Yamaji, Hiroshi Ueda

    Protein–protein interaction assays are important in various fields including molecular biology, diagnostics, and drug screening. We recently designed a novel protein–protein interaction assay, the firefly luminescent intermediate-based protein interaction assay (FlimPIA), that exploited the unique reaction mechanism of firefly luciferase (Fluc). Using two mutant Flucs, each impaired with one of the two half-reactions, namely adenylation and subsequent oxidative luminescent steps, FlimPIA detects the proximity of the two proteins tethered to the mutant Flucs. Here, we found that introducing a mutation into a residue in the hinge region (S440) of the mutant with lowered adenylation activity (‘Acceptor’ Fluc) further improved the response of FlimPIA by lowering the residual adenylation activity. Mutants with bulkier residues showed greater inhibition, probably due to increased steric hindrance at the adenylation conformation. As a result, the FlimPIA with S440 L acceptor showed the best signal/background ratio for the detection of rapamycin-induced FKBP12–FRB interactions.

    John Wiley and Sons Ltd, 2018年02月01日, Luminescence, 33 (1), 125 - 130, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Keita Mori, Hirotsugu Hamada, Takafumi Ogawa, Yuki Ohmuro-Matsuyama, Tomohisa Katsuda, Hideki Yamaji

    Transient gene expression allows a rapid production of diverse recombinant proteins in early-stage preclinical and clinical developments of biologics. Insect cells have proven to be an excellent platform for the production of functional recombinant proteins. In the present study, the production of an antibody Fab fragment by transient gene expression in lepidopteran insect cells was investigated. The DNA fragments encoding heavy-chain (Hc; Fd fragment) and light-chain (Lc) genes of an Fab fragment were individually cloned into the plasmid vector plHAneo, which contained the Bombyx mori actin promoter downstream of the B. mori nucleopolyhedrovirus (BmNPV) IE-1 transactivator and the BmNPV HR3 enhancer for high-level expression. Trichoplusia ni BTI-TN-5B1-4 (High Five) cells were co-transfected with the resultant plasmid vectors using linear polyethyleneimine. When the transfection efficiency was evaluated, a plasmid vector encoding an enhanced green fluorescent protein (EGFP) gene was also co-transfected. Transfection and culture conditions were optimized based on both the flow cytometry of the EGFP expression in transfected cells and the yield of the secreted Fab fragments determined by enzyme-linked immunosorbent assay (ELISA). Under optimal conditions, a yield of approximately 120 mg/L of Fab fragments was achieved in 5 days in a shake-flask culture. Transient gene expression in insect cells may offer a promising approach to the high-throughput production of recombinant proteins. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2017年08月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 124 (2), 221 - 226, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Yuki Ohmuro-Matsuyama, Keita Mori, Hirotsugu Hamada, Hiroshi Ueda, Hideki Yamaji

    Monoclonal antibodies and antibody fragments are used for diverse diagnostic and therapeutic applications. We have investigated the secretory production of Fab fragments from insect cells cotransfected with plasmid vectors carrying heavy- and light-chain genes. In the present study, to promote the formation of the disulfide bond between the heavy and light chains, some positively charged amino acid residues were introduced near the cysteine residue for the disulfide bond at the C-terminus of C-L, while some negatively charged amino acid residues were added near the cysteine residue for the disulfide bond at the C-terminus of C-H1. This electrostatic steering led to an increase in Fab secretions from insect cells.

    SPRINGER, 2017年06月, CYTOTECHNOLOGY, 69 (3), 469 - 475, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Production of Japanese encephalitis virus-like particles using insect cell expression systems

    Hideki Yamaji, Eiji Konishi

    2016年08月, Methods in Molecular Biology, 1404 (2), 365 - 375, 英語

    [招待有り]

    研究論文(学術雑誌)

  • Hideki Yamaji, Keita Mori, Hirotsugu Hamada, Yuki Ohmuro-Matsuyama, Tomohisa Katsuda

    ELSEVIER SCIENCE BV, 2016年07月, NEW BIOTECHNOLOGY, 33, S200 - S201, 英語

    研究論文(国際会議プロシーディングス)

  • Hideki Yamaji

    SPRINGER, 2016年06月, IN VITRO CELLULAR & DEVELOPMENTAL BIOLOGY-ANIMAL, 52 (Suppl 1), S10 - S10, 英語

    [招待有り]

    研究論文(国際会議プロシーディングス)

  • Makoto Kurihara, Yuki Ohmuro-Matsuyama, Keiichi Ayabe, Takahiro Yamashita, Hideki Yamaji, Hiroshi Ueda

    Detecting and assaying protein-protein interactions are significant research procedures in biology and biotechnology. We recently reported a novel assay to detect protein-protein interaction, i.e. firefly luminescent intermediate-based protein-protein interaction assay (FlimPIA) using two mutant firefly luciferases (Flucs), which complement each other's deficient half reaction. This assay detects neighboring of two mutant Flucs, namely a "Donor" that catalyzes the adenylation of firefly luciferin to produce a luciferyl-adenylate intermediate, and an "Acceptor" that catalyzes the subsequent light emitting reaction. However, its rather high background signal, derived from the remaining adenylation activity of the Acceptor, has limited its usefulness. To reduce this background signal, we introduced a mutation (R437K) into the hinge region of the Acceptor, while maintaining the oxidative activity. Interestingly, the signal/background (S/B) ratio of the assay was markedly improved by the addition of coenzyme A and reduction of the ATP concentration, probably due to reduced inhibition by dehydroluciferyl-adenylate formed during the catalysis and an increased ATP-based K-m value of the Acceptor, respectively. As a result, a significantly improved maximal S/B ratio from 2.5 to similar to 40 was attained, which promises wider use of the assay in in vitro diagnostics, drug discovery, and expanding our knowledge of various biological phenomena.

    WILEY-V C H VERLAG GMBH, 2016年01月, BIOTECHNOLOGY JOURNAL, 11 (1), 91 - 99, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Shinya Takada, Takafumi Ogawa, Kazusa Matsui, Tasuku Suzuki, Tomohisa Katsuda, Hideki Yamaji

    The generation of a recombinant baculovirus that displays antibody Fab fragments on the surface was investigated. A recombinant baculovirus was engineered so that the heavy chain (Hc; Fd fragment) of a mouse Fab fragment was expressed as a fusion to the N-terminus of baculovirus gp64, while the light chain of the Fab fragment was simultaneously expressed as a secretory protein. Following infection of Sf9 insect cells with the recombinant baculovirus, the culture supernatant was analyzed by enzyme-linked immunosorbent assay using antigen-coated microplates and either an anti-mouse IgG or an anti-gp64 antibody. A relatively strong signal was obtained in each case, showing antigen-binding activity in the culture supernatant. In western blot analysis of the culture supernatant using the anti-gp64 antibody, specific protein bands were detected at an electrophoretic mobility that coincided with the molecular weight of the Hc-gp64 fusion protein as well as that of gp64. Flow cytometry using a fluorescein isothiocyanate-conjugated antibody specific to mouse IgG successfully detected the Fab fragments on the surface of the Sf9 cells. These results suggest that immunologically functional antibody Fab fragments can be displayed on the surface of baculovirus particles, and that a fluorescence-activated cell sorter with a fluorescence-labeled antigen can isolate baculoviruses displaying specific Fab fragments. This successful baculovirus display of antibody Fab fragments may offer a novel approach for the efficient selection of specific antibodies.

    SPRINGER, 2015年08月, CYTOTECHNOLOGY, 67 (4), 741 - 747, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Luca Giannelli, Hiroyuki Yamada, Tomohisa Katsuda, Hideki Yamaji

    The green alga Haematococcus pluvialis, which accumulates astaxanthin at an optimal temperature of 20 degrees C, was cultivated under temperatures of 20 degrees C, 23.5 degrees C, 27 degrees C, and 30.5 degrees C, in order to assess the effects on algal metabolism during the growth phase. The culture growth rate declined with above-optimal increases in temperature, and the final maximum cell concentration at 30.5 degrees C reached only 35% of that attained at 20 degrees C. On the contrary, the biomass productivity was increased under all the high-temperature conditions, probably reflecting the metabolism switch from cell duplication to energy accumulation that is typically observed in algal cultures subjected to environmental stress. Moreover, an increase in the light-harvesting capability of the alga was observed by means of the total pigment balance and the photosynthesis-intensity (PI) curve measured under the different cultivation conditions. Cultures kept at higher temperatures were able to better harvest and utilize the impinging light due to photo-acclimation. Finally, the differences in the astaxanthin metabolism were elucidated by subjecting the cultures to nitrogen starvation at 20 degrees C and 27 degrees C. In the culture at 27 degrees C, a 1.4-fold increase in the astaxanthin productivity was observed when compared to that at 20 degrees C, and the latter required almost two-fold more energy for the astaxanthin production compared with the 27 degrees C culture. (C) 2014, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2015年03月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 119 (3), 345 - 350, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Luca Giannelli, Hideki Yamaji, Tomohisa Katsuda

    Vertical photobioreactors (PBR) with cylindrical cross section, namely air-lift reactors (ALR) and bubble column reactors (BCR), are often chosen both for bench-scale and industrial scale microalgal cultivation. It was common belief that ALR was the most favorable configuration in terms of light conversion effciency (LCE) and/or photosynthetic productivity than BCR because of the regular cyclic-flow pattern achieved inside the PBR. In the present study, we simulated the flow patterns in both ALR and BCR by means of computational fluid dynamics (CFD) and clarified the effects of such flow pattern on the LCE and productivity. Simulation results, obtained from the open-source CFD suite OpenFOAM, showed good agreement both for the flow velocity and the mixing time observed in the actual PBR using high-speed photography and conductivity pulse response, respectively. Subsequently, Lagrangian particle tracking was conducted on the simulation results to highlight the main fluid-flow patterns and to calculate the local-flashing-light frequency, which was necessary in order to estimate the overall light conversion effciency of the PBR. The BCR was characterized by a highly random fluid pattern with macroscopic, low-frequency circular loops while the ALR was characterized by numerous swirling flows localized inside the draft tube in addition to the main recirculation between the inner and outer portions of the tube. Finally, image analysis was used to correlate the numerical calculations with the light conversion effciencies attained in a Haematococcus pluvialis culture that had been illuminated with flashing light.

    SOC CHEMICAL ENG JAPAN, 2015年01月, JOURNAL OF CHEMICAL ENGINEERING OF JAPAN, 48 (1), 61 - 71, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hideki Yamaji, Tomohisa Katsuda, Hirotsugu Hamada, Keita Mori

    ELSEVIER SCIENCE BV, 2014年07月, NEW BIOTECHNOLOGY, 31, S186 - S186, 英語

    研究論文(国際会議プロシーディングス)

  • Hideki Yamaji

    Virus-like particles (VLPs) can be produced in recombinant protein production systems by expressing viral surface proteins that spontaneously assemble into particulate structures similar to authentic viral or subviral particles. VLPs serve as excellent platforms for the development of safe and effective vaccines and diagnostic antigens. Among various recombinant protein production systems, the baculovirus-insect cell system has been used extensively for the production of a wide variety of VLPs. This system is already employed for the manufacture of a licensed human papillomavirus-like particle vaccine. However, the baculovirus-insect cell system has several inherent limitations including contamination of VLPs with progeny baculovirus particles. Stably transformed insect cells have emerged as attractive alternatives to the baculovirus-insect cell system. Different types of VLPs, with or without an envelope and composed of either single or multiple structural proteins, have been produced in stably transformed insect cells. VLPs produced by stably transformed insect cells have successfully elicited immune responses in vivo. In some cases, the yield of VLPs attained with recombinant insect cells was comparable to, or higher than, that obtained by baculovirus-infected insect cells. Recombinant insect cells offer a promising approach to the development and production of VLPs.

    SPRINGER, 2014年03月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 98 (5), 1963 - 1970, 英語

    [査読有り][招待有り]

    研究論文(学術雑誌)

  • Hideki Yamaji, Masataka Nakamura, Miwa Kuwahara, Yusuke Takahashi, Tomohisa Katsuda, Eiji Konishi

    The production of Japanese encephalitis (JE) virus-like particles (VLPs) in stably transformed lepidopteran insect cells was investigated. The DNA fragment encoding the JE virus (JEV) prM signal peptide, the precursor (prM) of the viral membrane protein (M), and the envelope glycoprotein (E) was cloned into the plasmid vector pIHAbla. The pIHAbla contained the Bombyx mori actin promoter downstream of the B. mori nucleopolyhedrovirus (BmNPV) IE-1 transactivator and the BmNPV HR3 enhancer for high-level expression, together with a blasticidin resistance gene for use as a selectable marker. DNA encoding a form of prM with a pr/M cleavage site mutation was used to suppress the cell-fusion activity of VLPs. After transfection with the resultant plasmid, Trichoplusia ni BTI-TN-5B1-4 (High Five) cells were incubated with blasticidin, and cells resistant to the antibiotic were obtained. Western blot analysis and enzyme-linked immunosorbent assay of a culture supernatant showed that transfected High Five cells secreted an E antigen equivalent to the authentic JEV E. Sucrose density-gradient sedimentation analysis of the culture supernatant from recombinant High Five cells indicated that secreted E antigen molecules were produced in a particulate form. VLPs recovered from the supernatant successfully induced neutralizing antibodies in mice, particularly when adsorbed to alum adjuvant. High yields (a parts per thousand 30 mu g/ml) of E antigen were achieved in shake-flask cultures. These results indicate that recombinant insect cells may offer a novel approach for efficient VLP production.

    SPRINGER, 2013年02月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 97 (3), 1071 - 1079, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hideki Yamaji, Eiji Konishi

    Virus-like particles (VLPs) are composed of one or several recombinant viral surface proteins that spontaneously assemble into particulate structures without the incorporation of virus DNA or rna. The baculovirus-insect cell system has been used extensively for the production of recombinant virus proteins including VLPs. While the baculovirus-insect cell system directs the transient expression of recombinant proteins in a batch culture, stably transformed insect cells allow constitutive production. in our recent study, a secretory form of Japanese encephalitis (Je) VLPs was successfully produced by Trichoplusia ni BTi-Tn-5B1-4 (high Five) cells engineered to coexpress the Je virus (JEV) premembrane (PRM) and envelope (e) proteins. a higher yield of e protein was attained with recombinant high Five cells than with the baculovirus-insect cell system. This study demonstrated that recombinant insect cells offer a promising approach to the high-level production of VLPs for use as vaccines and diagnostic antigens. © 2013 Landes Bioscience.

    2013年, Bioengineered, 4 (6), 438 - 442, 英語

    [査読有り][招待有り]

    研究論文(学術雑誌)

  • YAMAJI Hideki, SEGAWA Maiko, NAKAMURA Masataka, KATSUDA Tomohisa, KUWAHARA Miwa, KONISHI Eiji

    The production of a secreted form of Japanese encephalitis (JE) virus-like particles (VLPs) using the baculovirus-insect cell system was investigated. A recombinant baculovirus that contained the JE virus (JEV) prM signal sequence and the genes encoding the precursor (prM) of the viral membrane protein (M) and the envelope glycoprotein (E) was constructed. Western blotting and enzyme-linked immunosorbent assay (ELISA) of the culture supernatant showed that Spodoptera frugiperda Sf9 cells infected with the recombinant baculovirus had secreted the E protein. Sucrose density-gradient sedimentation analysis of the culture supernatant suggested that secreted E antigen molecules were in a particulate form. Baculovirus-infected Sf9 cells produced more than a 10-fold higher yield of E antigen than that produced by previously reported recombinant CHO cells. Following infection with a recombinant baculovirus encoding a form of prM with a pr/M cleavage site mutation designed to suppress cell-fusion activity of E, Sf9 cells showed an E antigen yield comparable to a yield obtained with the baculovirus encoding the authentic form of prM. Baculovirus-infected Trichoplusia ni BIT-TN-5B1-4 (High Five) cells secreted less of the E antigen than Sf9 cells. Moreover, the Drosophila BiP signal sequence gave an E antigen yield comparable to the prM signal sequence, while the honeybee melittin signal sequence and the baculovirus gp64 signal sequence resulted in lower yields of the E antigen. These results provide information important to the development of VLP production processes using the baculovirus insect cell system. (C) 2012, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2012年12月, Journal of Bioscience and Bioengeering, 114 (6), 657 - 662, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hideki Yamaji, Miwa Kuwahara, Eiji Konishi

    ELSEVIER SCIENCE BV, 2012年09月, NEW BIOTECHNOLOGY, 29, S206 - S206, 英語

    研究論文(国際会議プロシーディングス)

  • Hiroyuki Sonoda, Yoichi Kumada, Tomohisa Katsuda, Hideki Yamaji

    We describe the secretory production of a mouse anti-bovine ribonuclease A single-chain variable fragment (scFv) fused with the Fc region of human IgG1 in stably transformed lepidopteran insect cells. Use of the insect-derived Bip and melittin signal peptides resulted in higher yields of the secreted scFv-Fc fusion protein than use of the baculovirus gp64 signal peptide. After cotransfection with an expression vector that contains the Bip signal sequence upstream of the DNA encoding the scFv-Fc and a selection vector that carries a neomycin resistance gene, Trichoplusia ni BT1-TN-5B1-4 (High Five) cells were cultured in the presence of G418. Colonies of resistant cells were obtained around 2 weeks after adding G418, and clonal cells were screened by enzyme-linked immunosorbent assay (ELISA) of the culture supernatant. The yield of the scFv-Fc protein secreted from the most productive clone was around 60 mg/l in a shake-flask culture. To improve the productivity, we investigated the effect of medium supplements of sodium butyrate (NaBu), dimethyl sulfoxide (DMSO), and sericin. Supplementing culture medium with sericin increased the scFv-Fc protein yield to 82 mg/l, but productivity was not increased by either NaBu or DMSO. These results indicate that the stably transformed insect cells may allow for the efficient production of scFv-Fc and other Fc fusion proteins. (C) 2012 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2012年08月, BIOCHEMICAL ENGINEERING JOURNAL, 67 (-), 77 - 82, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyoshi Miyahara, Ryou Nakashima, Masaki Inoue, Tomohisa Katsuda, Hideki Yamaji, Shigeo Katoh

    A silica-based medium with optimized pore size, pore volume, and coupling density was prepared for protein A affinity chromatography, to be used for industrial purification of IgG. Performance and durability of the silica-based medium were studied during repeated use and compared with other protein A media available on the market. The silica-based medium provided high dynamic binding capacities for human polyclonal IgG in the superficial liquid velocity range from 150 to 720?cm?h1 because of a high mass transfer coefficient and high mechanical strength. The dynamic binding capacity and IgG recovery were high and varied only slightly during 50150 cycles of repeated use. The release of protein A ligand was low.

    WILEY-BLACKWELL, 2012年01月, CHEMICAL ENGINEERING & TECHNOLOGY, 35 (1), 157 - 160, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Furuta, Takanori, Ogawa, Takafumi, Yamaji, Hideki

    2012年, Methods in Molecular Biology, 907, 英語

    [招待有り]

    研究論文(学術雑誌)

  • Tomohisa Katsuda, Hiroyuki Sonoda, Yoichi Kumada, Hideki Yamaji

    Escherichia coli is a host widely used in the industrial production of recombinant proteins. However, the expression of heterologous proteins in E. coli often encounters the formation of inclusion bodies, which are insoluble and nonfunctional protein aggregates. For the successful production of antibody fragments, which includes single-chain variable fragments (scFvs), we describe here the modification of linker, signal, and Shine-Dalgarno (SD) sequences, the coexpression of cytoplasmic and periplasmic chaperones, and a method for fed-batch cultivation with exponential feed. © 2012 Springer Science+Business Media, LLC.

    2012年, Methods in Molecular Biology, 907, 305 - 324, 英語

    [招待有り]

    研究論文(学術雑誌)

  • Production of antibody in insect cells

    YAMAJI Hideki

    2011年06月, Cell Engineering, Vol. 7, pp. 53-76, 英語

    [招待有り]

    研究論文(学術雑誌)

  • Hiroyuki Sonoda, Yoichi Kumada, Tomohisa Katsuda, Hideki Yamaji

    The effects of cytoplasmic and periplasmic chaperones on the secretory production of an anti-bovine ribonuclease A single-chain variable fragment (scFv) 3A21 in Escherichia coli were investigated. Co-expression of a cytoplasmic chaperone, GroEL/ES, DnaK/DnaJ/GrpE, trigger factor, or SecB with 3A21 scFv affected the proportions of antigen-binding activity in the cytoplasmic soluble fraction, the periplasmic fraction, and the extracellular medium, but there was no significant difference in the total activity compared to the control without chaperone co-expression. On the other hand, co-expression of a periplasmic chaperone, Skp or FkpA, with the exception of DsbC, greatly increased the binding activity in all the soluble fractions. Co-expression of both Skp and FkpA had no synergistic effect. Combinations of cytoplasmic and periplasmic chaperones decreased the productivity. In shake-flask cultures of cells co-expressing Skp or FkpA, considerable amounts of 3A21 scFv were detected in the extracellular medium by enzyme-linked immunosorbent assay (ELISA) and Western blot, and the extracellular production level of 3A21 scFv was calculated to be around 40mg/l. The binding activity of 3A21 scFv co-expressed with Skp was slightly higher than that with FkpA. These results indicate that the co-expression of periplasmic chaperones Skp and FkpA is extremely useful for the secretory production of scFvs in a culture medium using E. coli, but cytoplasmic chaperones and multiple-chaperone combinations may not be effective. Copyright (C) 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2011年04月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 111 (4), 465 - 470, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Sonoda, Yoichi Kumada, Tomohisa Katsuda, Hideki Yamaji

    We describe the soluble production of a mouse anti-bovine ribonuclease A single-chain variable fragment (scFv) fused with the Fc region of human IgG1 in Escherichia coli. Two production systems, secretory production using a pelB signal peptide and cytoplasmic production in a trxB/gor double mutant strain with an oxidizing cytoplasm, were investigated for efficient production of soluble and functional scFv-Fc fusion protein. Antigen-binding activity was observed in both systems but almost all of the scFv-Fc that was expressed formed insoluble aggregates. Hence, the co-expression of molecular chaperones was examined. Co-expression of GroEL/GroES showed a 4.6-fold increase in antigen-binding activity in the cytoplasmic production system but not in the secretory system. By contrast, the other two chaperones, DnaK/DnaJ/GrpE and trigger factor, had no effect in either production system. The protein solubility was also improved markedly by the co-expression of GroEL/GroES and approximately 70% of the 3A21 scFv-Fc protein was soluble. A practical productivity of more than 10 mg/L was achieved with a simple batch shake-flask culture. These results indicate that the E. coli cytoplasmic production system with oxidizing cytoplasm and molecular chaperones might be one of the choices for the soluble production of scFv-Fcs and other Fc fusion proteins. (c) 2010 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE BV, 2011年02月, BIOCHEMICAL ENGINEERING JOURNAL, 53 (3), 253 - 259, 英語

    [査読有り]

    研究論文(学術雑誌)

  • FURUTA Takanori, OGAWA Takafumi, KATSUDA Tomohisa, FUJI Ikuo, YAMAJI Hideki

    The production of an Fab fragment of the catalytic antibody 6D9 in lepidopteran insect cells infected with a recombinant baculovirus that contained both the heavy chain (Hc) and light chain (Lc) genes of the Fab fragment was investigated. Western blotting and enzyme-linked immunosorbent assay (ELISA) of culture supernatant showed that baculovirus-infected Trichoplusia ni BTI-TN-5B1-4 (High Five) cells secreted an Fab fragment that retained antigen-binding activity. Infection of High Five cells with a recombinant baculovirus, in which the Lc and Hc genes were located downstream of the baculovirus p10 and polyhedrin promoters, respectively, produced a higher Fab fragment yield than that obtained with a baculovirus in which the Hc and Lc genes were downstream of the p10 and polyhedrin promoters, respectively. Baculovirus-infected High Five cells secreted more of the Fab fragment than Spodoptera frugiperda Sf9 cells. Moreover, use of the baculovirus gp64 signal sequence upstream of the Lc and Hc genes resulted in greater yield of the secreted Fab fragment than use of the insect-derived BiP and melittin signal sequences. Consequently, the Fab fragment was obtained in a high yield (>600 mu g/ml) in a shake-flask culture of High Five cells infected at a multiplicity of infection (MOI) of 10 plaque-forming units (pfu)/cell with the recombinant baculovirus in which the Lc and Hc genes with the gp64 signal sequence were expressed under the control of the p10 and polyhedrin promoters, respectively. These results indicate that the baculovirus-insect cell system may allow efficient production of antibody Fab fragments. (C) 2010, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2010年11月, Journal of Bioscience and Bioengineering, 110 (5), 577 - 581, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Production of Japanese encephalitis virus-like particles in insect cell expression systems

    YAMAJI Hideki, NAGASUGA Takashi, TAKAHASHI Yusuke, NAKAMURA Masataka, KATSUDA Tomohisa, KUWAHARA Miwa, KONISHI Eiji

    2010年10月, The 13th Asia Pacific Confederation of Chemical Engineering Congress Conference Proceeding, 10640, 英語

    [査読有り]

    研究論文(国際会議プロシーディングス)

  • Naoya Morishita, Tomohisa Katsuda, Shuji Kubo, Akinobu Gotoh, Hideki Yamaji

    Adenovirus vector production by anchorage-independent 293 cells immobilized using porous biomass support particles (BSPs) was investigated in static and shake-flask cultures for efficient large-scale production of adenovirus vectors for gene therapy applications. The density of cells immobilized within BSPs was evaluated by measuring their WST-8 (2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt) reduction activity. In shake-flask culture, 293-F cells, which were adapted to serum-free suspension culture, were not successfully retained within reticulated polyvinyl formal (PVF) resin BSPs (2 x 2 x 2 mm cubes) with matrices of relatively small pores (pore diameter 60 mu m). When the BSPs were coated with a cationic polymer polyethyleneimine, a high cell density of more than 10(7) cells cm(-3)-BSP was achieved in both static and shake-flask cultures with regular replacement of the culture medium. After infection with an adenovirus vector carrying the enhanced green fluorescent protein gene (Ad EGFP), the specific Ad EGFP productivity of the immobilized cells was comparable to the maximal productivity of non-immobilized 293-F cells by maintaining favorable conditions in the culture environment.

    SPRINGER, 2010年08月, CYTOTECHNOLOGY, 62 (4), 293 - 300, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Sonoda, Yoichi Kumada, Tomohisa Katsuda, Hideki Yamaji

    The production of a single-chain variable fragment (scFv) antibody against bovine ribonuclease A in the cytoplasm of Escherichia coli trxB/gor double mutant was investigated. Previous reports have shown that the thioredoxin (Trx) protein fusion strategy is useful for the correct folding of scFvs and that the expression of functional scFvs is increased by co-expression of molecular chaperones. In the present study, we examined the effects of the combination of Trx fusion and molecular chaperone co-expression on the production of a functional scFv. A Trx-fused scFv was obtained in the oxidizing cytoplasm, and co-expression of GroELS and trigger factor had the greatest effect, resulting in a 2.8-fold increase in specific productivity. By contrast, the molecular chaperone DnaKJE had no effect. Moreover, co-expression of DnaKJE with GroELS negated the effects of GroELS. Trx-scFv was purified using a bovine ribonuclease A-coupled Sepharose column, and 2.7 mg/L of purified protein was obtained. Soluble Trx-scFv, expressed and purified as described above, exhibited pH-dependent binding similar to that of the parental full-length antibody. In addition, approximately 80% of the initial binding activity was retained after incubation at 37 degrees C for 2 weeks, indicating that the Trx-scFv fusion protein is quite stable. This strategy might be useful for the preparation of other recombinant scFvs. (C) 2009 Elsevier Inc. All rights reserved.

    ACADEMIC PRESS INC ELSEVIER SCIENCE, 2010年04月, PROTEIN EXPRESSION AND PURIFICATION, 70 (2), 248 - 253, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Beng Ti Tey, Kah Choon Yap, Hideki Yamaji, Abdul Manaf Ali, Wen Siang Tan

    SPRINGER, 2010年, ANIMAL CELL TECHNOLOGY: BASICS & APPLIED ASPECTS, 16, 73 - 76, 英語

    研究論文(国際会議プロシーディングス)

  • Hideki Yamaji, Yusuke Takahashi, Masataka Nakamura, Tomohisa Katsuda, Miwa Kuwahara, Eiji Konishi

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2009年11月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 108 (S1), S7 - S7, 英語

    研究論文(国際会議プロシーディングス)

  • Hiroyuki Yamamoto, Hideki Yamaji, Yuichiro Abe, Kazuo Harada, Danang Waluyo, Eiichiro Fukusaki, Akihiko Kondo, Hiromu Ohno, Hideki Fukuda

    Dimensionality reduction is an important technique for preprocessing of high-dimensional data. Because only one side of the original data is represented in a low-dimensional subspace, useful information may be lost. In the present study, novel dimensionality reduction methods were developed that are suitable for metabolome data, where observation varies with time. Metabolomics deal with this type of data, which are often obtained in microorganism fermentation processes. However, no dimensionality reduction method that utilizes information from the original data in a positive manner has been reported to date. The ordinary dimensionality reduction methods of principal component analysis (PCA), partial least squares (PLS), orthonormalized PLS (OPLS), and regularized Fisher discriminant analysis (RFDA) were extended by introducing differential penalties to the latent variables in each class. A nonlinear extension of this approach, using kernel methods, was also proposed in the form of kernel-smoothed PCA, PLS, OPLS, and FDA. Since all of these methods are formulated as generalized eigenvalue problems, the solutions can be computed easily. These methods were then applied to intracellular metabolite data of a xylose-fermenting yeast in ethanol fermentation. Visualization in the low-dimensional subspace suggests that smoothed PCA successfully preserves the information about the time course of observations during fermentation, and that RFDA can produce high separation among different strains. (c) 2009 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE BV, 2009年10月, CHEMOMETRICS AND INTELLIGENT LABORATORY SYSTEMS, 98 (2), 136 - 142, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Juan Huang, Na Jin, Tomohisa Katsuda, Hideki Fukuda, Hideki Yamaji

    Escherichia coli cells were efficiently immobilized in reticulated polyvinyl formal resin biomass support particles (BSPs) that had been simply autoclaved with a solution of a cationic polymer such as polyethyleneimine. When the immobilized E. coli cells containing aspartase were used as whole cell biocatalyst for L-aspartic acid production, they showed specific aspartase activity comparable to that of non-immobilized cells. (C) 2009 Elsevier B. V. All rights reserved.

    ELSEVIER SCIENCE SA, 2009年09月, BIOCHEMICAL ENGINEERING JOURNAL, 46 (1), 65 - 68, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Sonoda, Katsuya Daimon, Hideki Yamaji, Atsushi Sugimura

    The Vibrio proteolyticus aminopeptidase is synthesized as a preproprotein and then converted into an active enzyme by cleavage of the N-terminal propeptide. In recombinant Escherichia coli, however, the aminopeptidase is not processed correctly and the less-active form that has the N-terminal propeptide accumulates in the culture medium. Recently, we isolated a novel vibriolysin that was expressed as an active form in E. coli by random mutagenesis; this enzyme shows potential as a candidate enzyme for the processing of aminopeptidase. The E. coli cells were engineered to co-express the novel vibriolysin along with aminopeptidase. Co-expression of vibriolysin resulted in an approximately 13-fold increase in aminopeptidase activity, and a further increase was observed in the form lacking its C-terminal propeptide. The active aminopeptidase was purified from the culture supernatant including the recombinant vibriolysin by heat treatment and ion exchange and hydroxyapatite chromatography with high purity and 35% recovery rate. This purified aminopeptidase effectively converted methionyl-human growth hormone (Met-hGH) to hGH. Thus, this co-expression system provides an efficient method for producing active recombinant V. proteolyticus aminopeptidase.

    SPRINGER, 2009年08月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 84 (1), 191 - 198, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Kazutaka Ikeda, Yoichi Kumada, Tomohisa Katsuda, Hideki Yamaji, Shigeo Katoh

    A single-chain variable fragment (scFv) antibody against bisphenol A was refolded using an antigen (bisphenol A)-coupled column. The refolding efficiency was compared with that used in dialysis. The refolding efficiency of the antigen-coupled column was about 50-60%, which was much higher than with dialysis, due to a decrease in the concentration of the refolding molecules and to the suppression of the aggregate formation. (C) 2009 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2009年05月, BIOCHEMICAL ENGINEERING JOURNAL, 44 (2-3), 289 - 291, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Takuya Shishido, Naoya Kurata, Myung Eui Yoon, Tsutomu Tanaka, Hideki Yamaji, Hideki Fukuda, Akihiko Kondo

    Insect cell expression systems are widely used to produce active recombinant proteins. Here, we have developed a high-level expression vector containing a selectable marker for continuous production of recombinant proteins in insect cells. The plasmid, pXIHAbla, developed in this study, established a polyclonal cell line 8 days shorter than pXINSECT-DEST38 and pBmAneo. In addition, pXIHAbla exhibited an approximately fivefold higher average enhanced GFP expression level and approximately a twofold higher bionanocapsule secretion level than pXINSECT-DEST38. Using this plasmid, insect cells that highly express active proteins have been easily established.

    SPRINGER, 2009年05月, BIOTECHNOLOGY LETTERS, 31 (5), 623 - 627, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Kentaro Yamada, Naoya Morishita, Tomohisa Katsuda, Shuji Kubo, Akinobu Gotoh, Hideki Yamaji

    The use of low-multiplicity infection of 293 cells in static culture with regular medium replacement was investigated for efficient large-scale production of adenovirus vectors for gene therapy applications. An adenovirus vector carrying the enhanced green fluorescent protein gene (Ad EGFP) was used to infect 293-F cells at a low multiplicity of infection (MOI) of 0.00001-0.1 transductional unit (TU) cell(-1). The cells, which have the ability to grow in suspension, were incubated in T-flasks and the serum-free culture medium was replaced with fresh medium via centrifugation every 2 days. Because only a small proportion of cells were initially infected at low MOIs (< 1 TU cell(-1)), uninfected cells continued to grow until they were infected by progeny adenoviruses released from previously infected cells. When 293-F cells at a relatively low density of 1 x 10(5) cells cm(-3) were infected with Ad EGFP at a low MOI of 0.001 TU cell(-1), the vector yield was 2.7-fold higher than the maximum yield obtained with high-multiplicity infection (MOI = 10 TU cell(-1)) in batch culture. These results indicate that efficient adenovirus vector production using low MOIs is achieved by minimization of either nutrient depletion and/or accumulation of inhibitory metabolites in the culture medium.

    SPRINGER, 2009年04月, CYTOTECHNOLOGY, 59 (3), 153 - 160, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Yoichi Kumada, Yoshinobu Sakan, Hideyuki Kajihara, Mana Kihara, Yasufumi Kikuchi, Hideki Yamaji, Gi Hun Seong, Shigeo Katoh

    Single-chain Fv antibody (scFv) having 2 types of polypeptide linkers with or without rare codons, namely scFv (G(4)S)(3)(R) and scFv No.10 (with rare codons) and scFv (G(4)S)(3) and scFv No.10(NR) (without rare codon), were expressed under controllable conditions in batch and fed-batch fermentation, in order to compare volumetric productivity and specific productivity levels of scFvs as a soluble form. In batch fermentation, volumetric productivity levels of scFv (G(4)S)(3)(R) and scFv No.10, namely scFvs having the rare coclon linkers were 3-5 times higher than those of scFvs that had linkers without the rare codon. In fed-batch fermentation controlled by an exponential feeding system, the cell concentrations of the transformants increased with similar specific growth rates (0.1 h(-1)), while the specific productivity levels of scFvs with the rare codon linkers were 1.6 times higher than those of scFvs without the rare codon linkers. These results indicate that the presence of several rare codons in the gene of a polypeptide linker increases soluble amount of scFvs. This might be caused by a temporary decrease in translation speed at the position of the polypeptide linker allowing time for the folding of the V(H) domain and avoiding unfavorable interactions between amino acid residues at the unfolded V(H) and V(L) domains. Higher specific productivity levels of both scFv No. 10 and scFv No. 10(NR) than those of scFv (G(4)S)(3)(R) and (G(4)S)(3) might be caused by difference in stability of the polypeptide linkers on the basis of amino acid sequences. Thus, the rare codon linkers tested in this study will be considerably useful for large-scale production of soluble and active scFvs in fed-batch or continuous fermentations, in which high cell activity can be maintained. (C) 2008, The Society for Biotechnology, Japan. All rights reserved.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2009年01月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 107 (1), 73 - 77, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Mayu Fukui, Shinji Shiomi, Naoki Tanma, Tomohisa Katsuda, Naofumi Shiomi, Hideki Yamaji

    2009年, Journal of Biosceince and BIoengineering, 108 (1), S93 - S94, 日本語

    [査読有り]

    研究論文(国際会議プロシーディングス)

  • Hideki Yamaji, Toshitaka Manabe, Keizo Watakabe, Masaru Muraoka, Ikuo Fujii, Hideki Fukuda

    The production of an Fab fragment of the catalytic antibody 6D9 in stably transformed lepidopteran insect cells was investigated. On the basis of an expression vector that utilizes the Bombyx mori cytoplasmic actin promoter, from which foreign gene expression is stimulated with the B. mori nucleopolyhedrovirus (BmNPV) IE-1 transactivator and the BmNPV HR3 enhancer, two plasmid vectors were constructed which contain either a neomycin or a blasticidin resistance gene for use as a selectable marker. The genes encoding the heavy chain (Hc: Fd fragment) and light chain (Lc) of the 6D9 Fab fragment were inserted separately into the expression vectors. After cotransfection with the resulting plasmids to introduce the He and Lc genes and the two different antibiotic resistance genes, Trichoplusia ni BTI-TN-5B1-4 (High Five) cells were cultured in the presence of 6418 and blasticidin. Colonies of cells resistant to the antibiotics were obtained around 2 weeks after cotransfection. Western blotting and enzyme-linked immunosorbent assay (ELISA) of the cell culture supernatant suggested that the resistant cells stably secrete an Fab fragment which retains an antigen-binding activity. High yields of over 300 mu g/ml of Fab fragment were achieved in simple batch shake-flask culture of transfected insect cells. These results indicate that recombinant insect cells may offer a novel approach for efficient production of antibody molecules. (C) 2008 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2008年10月, BIOCHEMICAL ENGINEERING JOURNAL, 41 (3), 203 - 209, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Yuka Kobayashi, Tadashi Yamauchi, Tomohisa Katsuda, Hideki Yamaji, Shigeo Katoh

    Angiotensin-1 converting enzyme (ACE) plays important roles in the regulation of blood pressure, and ACE inhibitory peptides in food materials have attracted attention for their antihypertensive function. In this study, the function of amino acids in ACE inhibitory tripeptides was clarified.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2008年09月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 106 (3), 310 - 312, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Reza Ranjbar, Ryota Inoue, Tomohisa Katsuda, Hideki Yamaji, Shigeo Katoh

    In photobioreactors, photosynthetic microorganisms are exposed to certain light/dark cycles caused by light intensity distribution and mixing inside the photobioreactor. In this study Haematococcus pluvialis was cultivated in an airlift and a bubble column photobioreactor, and the cell growth and astaxanthin production were compared to clarify the effects of liquid circulation.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2008年08月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 106 (2), 204 - 207, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Shinji Hama, Sriappareddy Tamalampudi, Naoki Shindo, Takao Numata, Hideki Yamaji, Hideki Fukuda, Akihiko Kondo

    To develop a new approach for improving heterologous protein production in Aspergillus oryzae, we focused on the functional role of the N-terminal region of Rhizopus oryzae lipase (ROL). Several N-terminal deletion variants of ROL were expressed in A. oryzae. Interestingly, a segment of 28 amino acids from the C-terminal region of the propeptide (N28) was found to be critical for secretion of ROL into the culture medium. To further investigate the role of N28, the ROL secretory process was visualized in vivo using ROL-green fluorescent protein (GFP) fusion proteins. In cells producing ROL with N28, fluorescence observations showed that the fusion proteins are transported through endoplasmic reticulum (ER), Golgi, and cell wall, which is one of the typical secretory processes in a eukaryotic cell. Because the expression of the mature ROL-GFP fusion protein induced fluorescence accumulation without its translocation into the ER, N28 is considered to play a crucial role in protein transport. When N28 was inserted between the secretion signal and GFP, fluorescence observations showed that GFP, which is originally a cytoplasmic protein, was efficiently translocated into the ER of A. oryzae, resulting in an enhanced secretion of mature GFP after proteolytic cleavage of N28. These findings suggest that N28 facilitates protein translocation into ER and can be a promising candidate for improving heterologous protein production in A. oryzae.

    SPRINGER, 2008年07月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 79 (6), 1009 - 1018, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Hiroyuki Yamamoto, Hideki Yamaji, Eiichiro Fukusaki, Hiromu Ohno, Hideki Fukuda

    Multivariate regression analysis is one of the most important tools in metabolomics studies. For regression of high-dimensional data, partial least squares (PLS) has been widely used. Canonical correlation analysis (CCA) is a classic method of multivariate analysis; it has however rarely been applied to multivariate regression. In the present study, we applied PLS and regularized CCA (RCCA) to high-dimensional data where the number of variables (p) exceeds the number of observations (N), N << p. Using kernel CCA with linear kernel can drastically reduce the calculation time of RCCA. We applied these methods to gas chromatography-mass spectrometry (GC-MS) data, which were analyzed to resolve the problem of Japanese green tea ranking. To construct a quality-predictive model, the optimal number of latent variables in RCCA determined by leave-one-out cross-validation (LOOCV) was significantly fewer than in PLS. For metabolic fingerprinting, we successfully identified important metabolites for green tea grade classification using PLS and RCCA. (C) 2007 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2008年06月, BIOCHEMICAL ENGINEERING JOURNAL, 40 (2), 199 - 204, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Enzymatic biodiesel fuel production using whole-cell biocatalysts immobilized within biomass support particles

    Hideki Fukuda, Hideki Yamaji, Akihiko Kondo, Hideo Noda, Shinji Hama

    2007年09月, Journal of Biotechnology, 131S, S23 - S24, 英語

  • Juan Huang, Hideki Yamaji, Hideki Fukuda

    The technique of cell immobilization using porous biomass support particles (BSPs), which is attractive from the point of view of simplicity and convenience, relies on the inherent ability of adhesive cells, as a consequence of their growth, to form films around the support material or the ability of flocculent cells to create floes within the porous structure. In the present study, the immobilization of Escherichia coli cells using BSPs was investigated in shake-flask culture. The density of the cells immobilized within the BSPs was evaluated by measuring their intracellular lactate dehydrogenase (LDH) activity. Since E. coli K12 cells were not successfully retained within reticulated polyvinyl formal (PVF) resin BSPs with matrices of relatively small pores (pore diameter 60 pm), coating the surface of the BSPs with various polymers was examined as a way of promoting cell attachment. When positively charged polyamino acids such as poly-L-lysine, poly-L-arginine, poly-L-histidine, and poly-L-ornithine were adsorbed onto the particle surface, they were found to increase the immobilized cell density, while neutral and negatively charged polyamino acids including poly-L-asparagine and poly-L-glutamic acid were not effective. These results indicate that E. coli cells can be efficiently immobilized in PVF resin BSPs by electrostatic interaction between the negatively charged ions of the cell surface and the positively charged polymers adsorbed onto the BSP surface. A significantly high immobilized cell density was also achieved by coating the surface of the BSPs with the synthetic polymeric amine polyethyleneimine.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2007年08月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 104 (2), 98 - 103, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Takuya Shishido, Masaru Muraoka, Hideki Yamaji, Akihiko Kondo, Hideki Fukuda

    L particles, composed of the L protein of the hepatitis B virus surface antigen, are candidates for a specific gene and drug delivery system. We previously constructed stably transfected insect cells for L particle production. In this study, the cells were successfully immobilized within porous biomass support particles (BSPs) in shake-flask culture. The immobilized cells showed a high specific productivity, comparable to the maximum productivities in static and shake-flask cultures of nonimmobilized cells.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2007年06月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 103 (6), 572 - 574, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Shinji Hama, Hideki Yamaji, Takahiro Fukumizu, Takao Numata, Sriappareddy Tamalampudi, Akihiko Kondo, Hideo Noda, Hideki Fukuda

    A packed-bed reactor (PBR) system using fungus whole-cell biocatalyst was developed for biodiesel fuel production by plant oil methanolysis. Lipase-producing Rhizopus oryzae cells were immobilized within 6 mm x 6 mm x 3 mm cuboidal polyurethane foam biomass support particles (BSPs) during batch cultivation in a 20-1 air-lift bioreactor. Emulsification of the reaction mixture containing soybean oils and water improved the methanolysis reaction rate. Using a high flow rate for the reaction mixture in the PBR caused exfoliation of the immobilized cells from the BSPs, while the inefficient mixing of the reaction mixture at low flow rates allowed the BSPs to be covered with a hydrophilic layer of high methanol concentration, leading to a significant decrease in lipase activity. A high methyl ester content of over 90% was achieved at a flow rate of 251/h in the first cycle of repeated batch methanolysis and a high value of around 80% was maintained even after the tenth cycle. Comparison with methanolysis reaction in a shaken bottle suggested that the PBR enhances repeated batch methanolysis by protecting immobilized cells from physical damage and excess amounts of methanol. The process presented here is therefore considered to be promising for industrial biodiesel-fuel production. (c) 2007 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2007年06月, BIOCHEMICAL ENGINEERING JOURNAL, 34 (3), 273 - 278, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Yamaji, Hideki, Fukuda, Hideki

    2007年, Journal of Biotechnology, 131S, S54 - S54, 英語

    研究論文(国際会議プロシーディングス)

  • 高田 新也, 山地 秀樹, 福田 秀樹

    公益社団法人 化学工学会, 2007年, 化学工学会 研究発表講演要旨集, 2007, 373 - 373, 日本語

  • Hiroyuki Yamamoto, Keishi Hada, Hideki Yamaji, Tomohisa Katsuda, Hiromu Ohno, Hideki Fukuda

    The analysis of data from analytical equipment will be an important factor in the execution of metabolomics. Self-modeling curve resolution (SMCR) is one of the theoretical techniques of chemometrics and has recently been applied to the data of hyphenated chromatography techniques. Alternating least squares (ALS) is a classical SMCR method. In ALS, however, different solutions are produced depending on randomly chosen initial values. Simulation in the present study showed that the use of a normalized constraint in calculating ALS was effective in avoiding this problem. We also improved the ALS algorithm by adding a regularized term (regularized ALS: RALS). Independent component analysis (ICA) is a comparatively new method and has been discussed very actively by information science researchers, but has still been applied only in very few cases to curve resolution problems in chemometrics studies. We applied RALS with a normalized constraint and ICA to the HPLC-DAD data of Haematococcus pluvialis metabolites and obtained a high accuracy of peak detection, suggesting that these curve resolution methods are useful for identification of metabolites in metabolomics. (c) 2006 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2006年12月, BIOCHEMICAL ENGINEERING JOURNAL, 32 (3), 149 - 156, 英語

    [査読有り]

    研究論文(学術雑誌)

  • S Hama, S Tamalampudi, T Fukumizu, K Miura, H Yamaji, A Kondo, H Fukuda

    To identify the lipase responsible for the methanolysis activity of fungus whole-cell biocatalysts, the lipase localization of Rhizopus oryzae cells was determined. Western blot analysis showed that R. oryzae cells produce two types of lipase with different molecular masses of 34 and 31 kDa; the former (ROL34) was bound to the cell wall, whereas the latter (ROL31) was mainly bound to the cell membrane. It was found that cell immobilization within reticulated polyurethane foam biomass support particles strongly inhibits the secretion of membrane-bound lipase into the culture medium. An investigation of the relationship between ROL34 and ROL31 suggested that ROL31 originates from the cleavage of a 28-amino-acid residue at the N-terminus of ROL34. The addition of olive oil to the culture medium led to the retention of increased amounts of lipase within the cell. This phenomenon was further confirmed by an immunofluorescence labeling of hyphal cells. When cells were cultivated with various substrate-related compounds, such as olive oil and oleic acid, the intracellular methanolysis activity strongly correlated with the relative amounts of the membrane-bound lipase, which suggests that ROL31 localized in the membrane plays a crucial role in the methanolysis activity of R. oryzae cells.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2006年04月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 101 (4), 328 - 333, 英語

    [査読有り]

    研究論文(学術雑誌)

  • H Yamaji, T Manabe, A Kitaura, E Izumoto, H Fukuda

    To develop an efficient biological production process using the baculovirus-insect cell system, recombinant protein production was investigated in an immobilized cell culture with medium replacement. Sf9 insect cells were naturally entrapped within reticulated polyvinyl formal resin biomass support particles (BSPs; 2 mm x 2 mm x 2 mm; pore diameter 30-50 mu m) in a 2.5-1 stirred-tank bioreactor. The immobilized cells were grown to a density of over 107 cells/cm(3) -BSP, with the medium of 10% fetal bovine serum (FBS)-supplemented TNM-FH replaced at appropriate intervals, before infection with a recombinant baculovirus carrying the P-galactosidase gene. When serum-free TNM-FH was used instead of 10% FBS -supplemented TNM-FH for medium replenishment on and after post-infection day 1, the immobilized cells showed a high P-galactosidase yield comparable to that obtained in a culture using the serum-supplemented medium throughout. This finding indicates that immobilized insect cell culture allows not only intensification of cell culture but also recombinant protein production in protein-free basal media. (c) 2005 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2006年02月, BIOCHEMICAL ENGINEERING JOURNAL, 28 (1), 67 - 72, 英語

    [査読有り]

    研究論文(学術雑誌)

  • 黄 娟, 山地 秀樹, 福田 秀樹

    公益社団法人 化学工学会, 2006年, 化学工学会 研究発表講演要旨集, 2006, 1007 - 1007, 日本語

  • 沼田 崇男, 福水 崇裕, 濱 真司, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    公益社団法人 化学工学会, 2006年, 化学工学会 研究発表講演要旨集, 2006, 950 - 950, 日本語

  • Immobilized cell culture using different media for efficient production of biochemicals

    YAMAJI Hideki, NODA Hideo, FUKUDA Hideki

    2005年07月, 7th World Congress of Chemical Engineering Congress Manuscripts, 英語

    [査読有り]

    研究論文(国際会議プロシーディングス)

  • M Oda, M Kaieda, S Hama, H Yamaji, A Kondo, E Izumoto, H Fukuda

    For biodiesel-fuel production by methanolysis of plant oils, Rhizopus oryzae cells producing a 1,3-positional specificity lipase were cultured with polyurethane foam biomass support particles (BSPs) in a 201 air-lift bioreactor, and the cells immobilized within BSPs were used as whole-cell biocatalyst in repeated batch-cycle methanolysis reaction of soybean oil. The whole-cell biocatalyst had a higher durability in the methanolysis reaction when obtained from air-lift bioreactor cultivation than from shake-flask cultivation. Following repeated methanolysis reaction using the whole-cell biocatalyst, analysis of the reaction mixture composition indicated that monoglycerides (MGs) decreased and free fatty acids (FFAs) increased with increasing water content in the reaction mixture, and that MGs, diglycerides (DGs), and triglycerides (TGs) increased with increasing number of reaction cycles. The isomers of MGs and DGs generated during the 20th methanolysis reaction cycle consisted of 2-MGs and 1,2(2,3)-DGs, respectively. The hydrolytic activity of the whole-cell biocatalyst, on the other hand, was stable regardless of the number of reaction cycles. It was demonstrated thus that the whole cell biocatalyst promotes acyl migration of partial glycerides, and that the facilitatory effect is increased by increase in the water content of the reaction mixture but it is lost gradually with increasing number of reaction cycles. (C) 2004 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2005年03月, BIOCHEMICAL ENGINEERING JOURNAL, 23 (1), 45 - 51, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Cell cycle analysis of Chinese hamster ovary cells stimulated by phosphatidic acid in serum-free culture

    H Yamaji, K Sakai, T Joho, E Izumoto, H Fukuda

    When recombinant Chinese hamster ovary cells were treated with pertussis toxin or genistein, not only lysophosphatidic acid (LPA) but also phosphatidic acid (PA) failed to stimulate progression through the cell cycle in serum-free culture, suggesting that PA and LPA induce cell growth through the same signal transduction pathway. Cell cycle analysis also indicates that cell growth promoted by PA results in enhanced protein production.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2004年12月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 98 (6), 487 - 489, 英語

    [査読有り]

    研究論文(学術雑誌)

  • S Hama, H Yamaji, M Kaieda, M Oda, A Kondo, H Fukuda

    To stabilize the lipase activity of Rhizopus oryzae cells as whole-cell biocatalysts, the effect of cell membrane fatty acid composition on biodiesel-fuel production was investigated. The fatty acid composition of the cell membrane was easily controllable by addition of various fatty acids to the culture medium. Oleic or linoleic acid-enriched cells showed higher initial methanolysis activity than saturated fatty acid-enriched cells, among which palmitic acid-enriched cells exhibited significantly greater enzymatic stability than unsaturated fatty acid-enriched cells. It was assumed that fatty acids significantly affect the permeability and rigidity of the cell membrane, and that higher permeability and rigidity lead to increases in methanolysis activity and enzymatic stability, respectively. When the optimal fatty acid ratio of 0.67, indicated by R-f [=oleic acid/(oleic acid + palmitic acid)], was adopted for repeated methanolysis reactions, both methanolysis activity and enzymatic stability were maintained at significantly elevated levels, with methyl ester content of around 55% even in the 10th batch cycles. (C) 2004 Elsevier B.V. All rights reserved.

    ELSEVIER SCIENCE SA, 2004年10月, BIOCHEMICAL ENGINEERING JOURNAL, 21 (2), 155 - 160, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Efficient protein production by insect cells immobilized within porous support particles

    MANABE Toshitaka, KITAURA Akinori, YAMAJI Hideki, FUKUDA Hideki

    2004年, The 10th APCChE Congress Conference Proceedings, 3P-01-086, 英語

    [査読有り]

    研究論文(国際会議プロシーディングス)

  • Comparative study on delivery of phosphatidic acid to serum-free culture of Chinese hamster ovary cells

    C Hayashi, K Sakai, H Yamaji, H Fukuda

    Liposomes containing phosphatidic acid (PA) and inclusion complexes of PA with methyl-P-cyclodextrin (CD) were compared with PA dispersion prepared with a nonionic surfactant, Tween 80, for their effect on the growth of Chinese hamster ovary (CHO) cells in serum-free culture. All of the types of PA preparation were capable of promoting cell growth to almost the same high extent, indicating that the efficacy of PA in stimulating cellular growth may not depend on the preparation method.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2003年08月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 96 (2), 196 - 198, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Production of protein kinase C-δ by the baculovirus-insect cell system in serumsupplemented and serum-free media

    YAMAJI Hideki, HIRAKAWA Daigo, TAGAI Shin-ichi, FUKUDA Hideki

    When rat protein kinase C-delta (PKC-delta) was produced by Sf9 cells infected with a recombinant baculovirus in shake-flask culture using a serum-containing medium, the intracellular PKC-delta content decreased in the late period while the extracellular PKC-delta markedly increased. During the late period of serum-free culture, the extracellular PKC-delta level considerably declined, but the addition of a protease inhibitor, leupeptin, prevented the reduction in PKC-delta production.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2003年02月, Journal of Bioscience and Bioengineering, 95 (2), 185 - 187, 英語

    [査読有り]

    研究論文(学術雑誌)

  • N Nishikawa, H Yamaji, H Fukuda

    The production of beta-galactosidase by Sf9 cells infected with recombinant Autographa californica nucleopolyhedrovirus ( AcNPV) was investigated in shake-flask culture using two serum-free basal media: Grace's medium and TNM-FH ( Grace's medium supplemented with lactalbumin hydrolysate and yeast extract). At the time of infection, cells grown in serum-supplemented TNM-FH were transferred into fresh basal media without adaptation. The absence of serum depressed the beta-galactosidase yield considerably in Grace's medium, but to a much lesser extent in TNM-FH, where it reached around 2/3 of the level obtained in TNM-FH supplemented with 10% fetal bovine serum (FBS). While both lactalbumin hydrolysate and yeast extract promoted beta-galactosidase production, their removal by medium replacement on post-infection day 1 gave a beta-galactosidase yield nearly equal to that obtained in their continuous presence. Supplementation of basal media with phosphatidic acid (PA) from egg yolk lecithin, which has been shown to enhance cell growth and recombinant protein production in serum-free culture of Chinese hamster ovary (CHO) cells, was also effective in increasing beta-galactosidase yield. Elevating the multiplicity of infection (MOI) from 2 to 10 plaque-forming units per cell (pfu/cell) also resulted in an increase in product yield. These results provide information important to the development of cost-effective serum-free culture technology for use in large-scale production of recombinant proteins by the baculovirus - insect cell system.

    KLUWER ACADEMIC PUBL, 2003年, CYTOTECHNOLOGY, 43 (1-3), 3 - 10, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Effects of phosphatidic acid on recombinant protein production by Chinese hamster ovary cells in serum-free culture

    K Sakai, T Matsunaga, C Hayashi, H Yamaji, H Fukuda

    When recombinant Chinese hamster ovary (CHO) cells (ATCC CRL-8200) producing human interferon-gamma (hIFN-gamma) were incubated with exogenously supplied phosphatidic acid (PA) from egg yolk lecithin in a serum-free medium, PA induced increases in both the density of viable cells and concentration of hIFN-gamma secreted into the medium. Dispersion of PA with a non-ionic surfactant, Tween 80, further enhanced both cell growth and hIFN-gamma production. Replacement of the culture medium containing PA by fresh medium without PA in the course of a static culture did not influence cell growth indicating that PA is required to be continuously present in a serum-free medium to stimulate cell growth. Using a fresh medium containing PA for replacement resulted in significant enhancement of both cell density and hIFN-gamma yield. These results suggest that PA is a promising constituent of low-protein serum-free media for the effective production of recombinant proteins. (C) 2002 Elsevier Science B.V All rights reserved.

    ELSEVIER SCIENCE SA, 2002年03月, BIOCHEMICAL ENGINEERING JOURNAL, 10 (2), 85 - 92, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Dialysis perfusion culture of animal cells for the production of biologicals

    Yamaji, H, Amos, B, Emery, AN, Al-Rubeai, M, Shirahata, S, Teruya, K, Katakura, Y

    2002年, Animal Cell Technology : Basic & Applied Aspects, Vol 12

    [査読有り]

    研究論文(学術雑誌)

  • Use of nonionic surfactants for effective supply of phosphatidic acid in serum-free culture of Chinese hamster ovary cells

    K Sakai, C Hayashi, H Yamaji, H Fukuda

    We have previously shown [Sakai et al., J. Biosci. Bioeng., 88, 306-309 (1999)] that exogenously supplied phosphatidic acid (PA) and lysophosphatidic acid (LPA) promoted the growth of Chinese hamster ovary (CHO) cells in serum-free culture. However, the direct addition of high concentrations of these phospholipids alone to the culture medium resulted in the formation of precipitates. We therefore examined the use of two nonionic surfactants, Tween 80 and Pluronic F-68, as a means of supplying PA more effectively to CHO cells in a serum-free culture. A clear dispersion of PA from egg yolk lecithin that could be successfully sterile-filtered was obtained by using Tween 80 or Pluronic F-68. When PA prepared with either of the surfactants was added to serum-free media, precipitation was noticeably reduced. Furthermore, the growth-promoting activity of PA was considerably enhanced by the presence of the surfactants. Since Tween 80 and Pluronic F-68 themselves possessed no growth-stimulating property, it was suggested that the enhanced growth-promoting activity results from the improved availability of PA to the cells. The use of Tween 80 with PA analogues having saturated acyl chains also accelerated cell growth, whereas these PAs showed little growth-promoting activity, due to their poor water-solubility, when added alone.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2001年09月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 92 (3), 256 - 261, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Miyagawa, S, Nakagawa, M, Ito, Y, Yamaji, H, Fukuda, H, Katoh, S

    2001年, Journal of Chemical Engineering of Japan, 34 (1)

    [査読有り]

    研究論文(学術雑誌)

  • Production of recombinant protein by baculovirus-infected insect cells in immobilized culture using porous biomass support particles

    H Yamaji, SI Tagai, K Sakai, E Izumoto, H Fukuda

    Immobilization of insect cells using porous biomass support particles (BSPs) and production of a recombinant protein by the immobilized cells after infection with a baculovirus were investigated in a shake-flask culture. Sf9 cells were passively immobilized in reticulated polyvinyl formal (PVF) resin BSPs (2 x 2 x 2 mm cubes) with matrices of 60 mu m mean pore diameter in situ in shake-flasks. The cell density in the BSPs was over 5 x 10(7) cells/cm(3)-BSP in cultures with regular replacement of the culture medium, as estimated by 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. After infection with a recombinant baculovirus carrying the beta-galactosidase gene, immobilized cells within the BSPs showed a high specific productivity, comparable to the maximum productivity in shake-flask cultures of non-immobilized cells, as long as nutrients in the medium were not depleted. Even when immobilized cells at a high density of 5 x 107 cells/cm(3)-BSP were infected with the baculovirus, efficient beta-galactosidase production with a high specific productivity was possible by replacing the medium at appropriate intervals to avoid nutrient depletion.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 2000年01月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 89 (1), 12 - 17, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Effects of phospholipids on growth of Chinese hamster ovary cells in serum-free media

    K Sakai, T Matsunaga, H Yamaji, H Fukuda

    The effects of phosphatidic acid (PA) and lysophosphatidic acid (LPA) dispersed in serum-free media on the growth of Chinese hamster ovary (CHO) cells were investigated. After cells were incubated in serum-free media supplemented with PA or LPA for 3 d, the cellular growth was evaluated by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Supplementing PA from egg yolk lecithin to basal synthetic media such as alpha-MEM (minimum essential medium, alpha modification) markedly promoted the growth of CHO cells. PA from egg yolk lecithin also enhanced the cell growth in a serum-free medium containing insulin and transferrin. When PA analogues with different acyl chains of C14-18 were compared, PA with unsaturated acyl chains, dioleoyl-PA (C18 : 1), was most effective far stimulating the growth of CHO cells. Similar results were obtained when LPA was examined. These results suggest that PA and LPA, especially those with unsaturated acyl chains, are promising growth-promoting supplements for use as constituents in a low-protein serum-free medium.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 1999年09月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 88 (3), 306 - 309, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Optimal production of recombinant protein by the baculovirus-insect cell system in shake-flask culture with medium replacement

    H Yamaji, S Tagai, H Fukuda

    Sf9 insect cells infected with a recombinant baculovirus expressing beta-galactosidase and suspended in fresh medium (TNM-FH supplemented with 10% fetal bovine serum) at the time of infection were cultured in shake flasks at various combinations of initial cell density and multiplicity of infection (MOI). The effects of cell density and MOI on beta-galactosidase production were quantitatively analyzed by plotting the beta-galactosidase yield against the time integral of the viable cell density from the time of infection to the time when the beta-galactosidase production reached a plateau. The beta-galactosidase yield had a maximum value at a viable cell density time integral of approximately 8 x 10(6) cells.d/cm(3) for each MOI used in a range from 0.01 to 10 plaque-forming units per cell (pfu/cell). Since glucose and fructose were exhausted when the culture reached 8 x 106 cells.d/cm(3), it was concluded that protein production in a high-cell-density culture was limited by nutrient depletion in the culture medium, and hence the nutritional capacity of the medium was able to be determined as the viable cell density time integral at which the maximum product yield was attained. In cultures infected at a low MOI (less than or equal to 1 pfu/cell), the specific productivity, and thereby the yield, of beta-galactosidase declined with decreasing MOI due to the reduction in the proportion of initially infected cells. These results indicate that production of a recombinant protein in a culture with medium replacement at the time of infection can be optimized if the cells are infected at a high MOI (greater than or equal to 1 pfu/cell) and at a cell density such that the viable cell density time integral reaches the nutritional capacity just as the protein production is completed.

    SOC BIOSCIENCE BIOENGINEERING JAPAN, 1999年05月, JOURNAL OF BIOSCIENCE AND BIOENGINEERING, 87 (5), 636 - 641, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Growth stimulation of CHO cells by phospholipids in serum-free culture

    Sakai, K, Matsunaga, T, Yamaji, H, Fukuda, H, Ikura, K, Nagao, M, Masuda, S, Sasaki, R

    1999年, Animal Cell Technology: Challenges For the 21st Century

    [査読有り]

    研究論文(学術雑誌)

  • Hideki Yamaji, Masato Nakagawa, Kanji Tomioka, Akihiko Kondo, Hideki Fukuda

    For spectrophotometric liposome immunoassay of a specific antibody, antigen (cytochrome c)-coupled liposomes containing the substrate glucose-6-phosphate (G6P) as a marker were prepared from 1,2-di(3RS,7R,11R-phytanyl)-sn-glycero-3-phosphocholine (DPhyPC). DPhyPC liposomes showed good stability against G6P leakage, which was below 8% after incubation for 70 d at 4°C. When cytochrome c-coupled liposomes were incubated with anti-cytochrome c antibody and complement, after which glucose-6-phosphate dehydrogenase and NAD+ were added, the absorbance at 340 nm increased with the amount of anti-cytochrome c antibody added.

    Elsevier Sci B.V., 1997年, Journal of Fermentation and Bioengineering, 83 (6), 596 - 598, 英語

    [査読有り]

    研究論文(学術雑誌)

  • Continuous IgG production by myeloma cells immobilized within porous support particles

    H Yamaji, H Fukuda

    IgG production kinetics of mouse myeloma MPC-11 cells immobilized within porous polyvinyl formal resin biomass support particles (BSPs) was evaluated by monitoring of lactate dehydrogenase activities and IgG concentration in a perfusion culture of the cells. Cells immobilized within the BSPs could stably maintain a high specific IgG production rate, comparable to the maximum production rate in a static culture of non-immobilized MPC-11 cells, over a long period.

    SOC FERMENTATION BIOENGINEERING, JAPAN, 1997年, JOURNAL OF FERMENTATION AND BIOENGINEERING, 83 (5), 489 - 491, 英語

    [査読有り]

    研究論文(学術雑誌)

  • GROWTH-KINETICS OF ANIMAL-CELLS IMMOBILIZED WITHIN POROUS SUPPORT PARTICLES IN A PERFUSION CULTURE

    H YAMAJI, H FUKUDA

    The growth kinetics of anchorage-independent animal cells [mouse myeloma MPC-11 (ATCC CCL 167)] immobilized within porous polyvinyl formal resin biomass support particles (BSPs; 3 x 3 x 3 mm cubes; mean pore diameter, 60 mu m) was analyzed by measuring intracellular and extracellular lactate dehydrogenase (LDH) activities in a perfusion culture. First, the intracellular LDH content of cells immobilized within the BSPs was assayed in a shake-flask culture and found to remain almost comparable to that of nonimmobilized cells in the exponential growth phase under static culture. Then, cells inoculated in the BSPs were grown in a continuous stirred-tank bioreactor. Using the LDH content of non-immobilized cells, the density of immobilized cells and the numbers of leaked and dead cells were evaluated, respectively, by the intracellular LDH activity of immobilized and leaked cells and the LDH activity in cell-free culture supernatant. In the initial period, immobilized cells exhibited exponential growth at a constant apparent specific growth rate; however, the actual specific growth rate, which takes into consideration cell death and cell leakage, decreased significantly. In the stationary phase, the actual specific growth rate maintained a constant but markedly lower value than during exponential growth.

    SPRINGER VERLAG, 1994年12月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 42 (4), 531 - 535, 英語

    [査読有り]

    研究論文(学術雑誌)

  • GROWTH AND DEATH BEHAVIOR OF ANCHORAGE-INDEPENDENT ANIMAL-CELLS IMMOBILIZED WITHIN POROUS SUPPORT MATRICES

    H YAMAJI, H FUKUDA

    Growth and death of anchorage-independent animal cells entrapped within porous biomass support particles (BSPs) in static or shake-flask cultures were evaluated by comparison of enzyme activity with non-immobilized cells grown under static culture using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and release of lactate dehydrogenase into the culture medium. Mouse myeloma MPC-11 (ATCC CCL 167) cells inoculated within porous polyvinyl formal resin BSPs (3 x 3 x 3 or 2 x 2 x 2 mm; mean pore diameter, 60-mu-m) grew exponentially at a specific growth rate comparable to that of non-immobilized cells in the initial period of incubation. Entrapped cells then reached the stationary phase with a cell density over 10(7) cells/cm3 BSP. The death rate of entrapped cells increased in response to the rise in viable cell density in the BSPs. Observation of viable cell distribution within the BSPs using MTT staining indicated that the cells concentrated within a thin outer shell of the BSPs with time. After the immobilized cells reached the stationary phase, penetration of cells into the outer shell ceased and heterogeneous distribution of cell density occurred in the viable cell layer in the shake-flask culture.

    SPRINGER VERLAG, 1992年05月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 37 (2), 244 - 251, 英語

    [査読有り]

    研究論文(学術雑誌)

  • A NOVEL BIOREACTOR PROCESS FOR ANCHORAGE-INDEPENDENT ANIMAL-CELLS IMMOBILIZED WITHIN POROUS BIOMASS SUPPORT PARTICLES

    H YAMAJI, H FUKUDA

    KLUWER ACADEMIC PUBL, 1992年, ANIMAL CELL TECHNOLOGY : BASIC & APPLIED ASPECTS, VOL 4, 239 - 244, 英語

    [査読有り]

    研究論文(国際会議プロシーディングス)

  • LONG-TERM CULTIVATION OF ANCHORAGE-INDEPENDENT ANIMAL-CELLS IMMOBILIZED WITHIN RETICULATED BIOMASS SUPPORT PARTICLES IN A CIRCULATING BED FERMENTER

    H YAMAJI, H FUKUDA

    Long-term cultivation of anchorage-independent animal cells immobilized within porous biomass support particles (BSPs) using a gas-stirred circulating bed fermentor (CBF) was investigated. Inoculation of mouse myeloma MPC-11 (ATCC CCL 167) cells into reticulated polyvinyl formal resin BSPs (3 x 3 x 3 mm; mean pore diameter, 60-mu-m; porosity, 0.88) and the repeated batch culture of inoculated cells were performed under gentle circulation of BSPs, induced by sparging air from the base of the fermentor. The glucose uptake rate of cells decreased in the initial period just after the start of circulation, since a relatively large number of cells leaked from the BSPs. After that period, the uptake rate gradually increased and the leakage of cells diminished. In the meantime, when inoculated cells were incubated statically by introducing air into the upper part of the fermentor for the initial several days before circulating the BSPs, glucose consumption became very rapid and cell density in the BSPs reached at least 10(7) cells/cm3 BSP. Thus, a long-term cultivation without significant leakage of cells and with high cell density in BSPs was successfully achieved in the CBF-BSP system.

    SPRINGER VERLAG, 1991年03月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 34 (6), 730 - 734, 英語

    [査読有り]

    研究論文(学術雑誌)

  • KISHIMURA, M, YAMAJI, H, FUKUDA, H, TERASHIMA, M, KATOH, S, SADA, E, TANIGUCHI, H

    1990年, Annals of the New York Academy of Sciences, 613

    [査読有り]

    研究論文(学術雑誌)

  • IMMOBILIZATION OF ANCHORAGE-INDEPENDENT ANIMAL-CELLS USING RETICULATED POLVINYL FORMAL RESIN BIOMASS SUPPORT PARTICLES

    H YAMAJI, H FUKUDA, Y NOJIMA, C WEBB

    SPRINGER VERLAG, 1989年06月, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 30 (6), 609 - 613, 英語

    [査読有り]

    研究論文(学術雑誌)

  • KISHIMURA, M, YAMAJI, H, FUKUDA, H, TERASHIMA, M, KATOH, S, SADA, E

    1989年, Journal of Fermentation and Bioengineering, 68 (6)

    [査読有り]

    研究論文(学術雑誌)

  • Yuki Ohmuro-Matsuyama, Keiko Gomi, Takuya Shimoda, Hideki Yamaji, Hiroshi Ueda

    2021年11月, Frontiers in Bioengineering and Biotechnology, 9, 778120, 英語

    [査読有り]

    研究論文(学術雑誌)

MISC

  • 昆虫細胞を用いた有用物質生産

    山地 秀樹

    日本生物工学会, 2016年02月, 生物工学, 94 (2), 76 - 80, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • 1P-083 酵素改良と反応条件至適化による相互作用検出系FlimPIAの高機能化(酵素学,酵素工学,一般講演)

    上田 宏, 栗原 誠, 山下 貴宏, 綾部 圭一, 山地 秀樹, 大室 有紀

    日本生物工学会, 2014年, 日本生物工学会大会講演要旨集, 66, 38 - 38, 日本語

  • 3P-160 シトクロムP450発現大腸菌の増殖と酵素活性に及ぼす培養条件の影響(生物化学工学,一般講演)

    森中 涼平, 山地 秀樹, 今石 浩正

    日本生物工学会, 2013年, 日本生物工学会大会講演要旨集, 65, 228 - 228, 日本語

  • 昆虫細胞を用いたウイルス様粒子の生産

    山地 秀樹

    2012年10月, PHARM TECH JAPAN, 28 (12), 2503 - 2508, 日本語

    [招待有り]

    記事・総説・解説・論説等(商業誌、新聞、ウェブメディア)

  • 3Hp25 Computational fluid dynamics for enhancing the light distribution in a photobioreactor grown culture of Hematococcus pluvialis :

    Giannelli Luca, Wada Sotaro, Yamaji Hideki, Katsuda Tomohisa

    日本生物工学会, 2012年, 日本生物工学会大会講演要旨集, 64, 157 - 157, 英語

  • 2Bp21 組換え昆虫細胞によるscFv-Fc融合タンパク質の生産(バイオプロセス/センサー,計測工学/セル&ティッシュエンジニアリング,一般講演)

    薗田 啓之, 熊田 陽一, 勝田 知尚, 山地 秀樹

    日本生物工学会, 2012年, 日本生物工学会大会講演要旨集, 64, 32 - 32, 日本語

  • 細胞はどこまで大きくなれる?

    山地 秀樹

    2011年03月, 化学工学, Vol. 75, No. 3, p. 163, 日本語

    記事・総説・解説・論説等(学術雑誌)

  • 組換え昆虫細胞によるウイルス様粒子の連続生産システムの構築

    山地 秀樹

    2009年06月, 平成19年度~平成20年度科学研究費補助金 (基盤研究 (C)) 研究成果報告書, 日本語

    その他

  • 昆虫細胞を用いた高発現システムによるタンパク質生産

    山地 秀樹

    2005年08月, 化学と生物, 43 (8), 491 - 492, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • 遺伝子治療用アデノウイルスベクター

    山地 秀樹

    2005年02月, 生物工学, 83 (2), 91 - 91, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • 植物油製造過程で発生する脱酸廃棄物からバイオ燃料の実用化生産

    福田 秀樹, 近藤 昭彦, 山地 秀樹

    2005年, 平成16年度 地域新生コンソーシアム研究開発事業成果報告書, 日本語

    その他

  • 昆虫細胞のバイオリアクター技術

    山地 秀樹, 福田 秀樹

    2004年12月, 生物工学, 82 (12), 589 - 591, 日本語

    [招待有り]

  • 昆虫細胞とバキュウロウイルスのアプリケーション 特集によせて

    山地 秀樹, 福田 秀樹

    2004年12月, 生物工学, 82 (12), 579 - 579, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • プロテオーム解析による新規細胞培養技術の開発

    山地 秀樹

    2004年, 平成14年度~平成15年度科学研究費補助金(基礎研究(C)(2))研究成果報告書, 日本語

    その他

  • 昆虫細胞の高密度培養とタンパク質生産

    山地 秀樹

    2003年02月, バイオサイエンスとインダストリー, 61 (2), 99 - 102, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • バイオ人工腱の生体力学的特性に及ぼす単軸方向の周期的な負荷の影響

    山地 秀樹

    2003年, (社)日本生物工学会 セル&ティッシュエンジニアリング研究部会編 「再生医療実用化に向けた生物工学研究 ー 英米および国内生物工学者の活動 ー, 9の11, 日本語

    その他

  • 遺伝子治療のためのレトロウイルスベクターの生産

    山地 秀樹

    2000年12月, 化学工学, 64 (12), 677 - 677

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • 多孔性粒子を用いる細胞の固定化

    山地 秀樹

    1999年12月, 化学工学, 63 (12), 682 - 683, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

  • 昆虫細胞-バキュロウイルスは究極のタンパク質生産系になりうるか?

    山地 秀樹

    1997年09月, 生物工学, 75 (5), 391 - 391, 日本語

    [招待有り]

    記事・総説・解説・論説等(学術雑誌)

書籍等出版物

  • バイオリアクターのスケールアップと物質生産事例集 (第7章2節「昆虫細胞を用いた組換えタンパク質の生産」(pp. 237–243) を分担執筆)

    山地秀樹

    分担執筆, 第7章2節 (pp. 237–243), 技術情報協会, 2021年04月

  • 医薬品モダリティの特許戦略と技術開発動向 (第2章7節「昆虫細胞を用いたバイオ医薬品の生産技術」(pp. 135–142) を分担執筆)

    山地 秀樹

    分担執筆, 第2章7節 (pp. 135–142), 技術情報協会, 2019年05月

  • バイオロジクスの開発と品質・安全性確保 上巻 (監修:早川堯夫),第1部第1章第1節第4項「昆虫細胞を用いたバイオ医薬品の生産」(pp. 74–80) を分担執筆

    山地 秀樹

    分担執筆, (株) エル・アイ・シー, 2018年09月, 日本語, ISBN: 9784900487567

    学術書

  • 動物細胞培養の手法と細胞死・増殖不良・細胞変異を防止する技術 (第2章第1節[2]「動物細胞の無血清培養におけるリン脂質の利用」(pp. 32–36) を分担執筆)

    山地 秀樹

    分担執筆, 技術情報協会, 2014年04月, 英語

    学術書

講演・口頭発表等

  • Adsorption of immunoglobulin G in protein A resins studied by confocal laser scanning microscopy taking account of the optical hindrance inside a particle

    Tomohisa Katsuda, Tomoki Yamaguchi, Naruaki Inoue, Yuki Kase, Kou Ota, Hideki Yamaji

    18th Asian Pacific Confederation of Chemical Engineering Congress (APCChE 2019), 2019年09月, 英語

    ポスター発表

  • Production of influenza virus proteins in recombinant insect cells

    Takuya Matsuda, Toshikazu Tanijima, Kyoko Masumi-Koizumi, Tomohisa Katsuda, Hideki Yamaji

    18th Asian Pacific Confederation of Chemical Engineering Congress (APCChE 2019), 2019年09月, 英語

    ポスター発表

  • 微細藻の細胞分裂に及ぼす培養温度・光強度の周期的制御の影響

    勝田 知尚, 森本 高章, 桑井 仁葵, 山地 秀樹

    第71回日本生物工学会大会, 2019年09月, 日本語

    口頭発表(一般)

  • 組換え昆虫細胞によるインフルエンザウイルス抗原タンパク質の生産

    松田 拓也, 増見 恭子, 勝田 知尚, 山地 秀樹

    第32回日本動物細胞工学会2019年度大会 (JAACT2019), 2019年07月, 日本語

    ポスター発表

  • Production of influenza virus-like particles in insect cells

    Hideki Yamaji, Takuya Matsuda, Toshikazu Tanijima, Kyoko Masumi-Koizumi, Tomohisa Katsuda

    26th ESACT (the European Society for Animal Cell Technology) Meeting (ESACT 2019), 2019年05月, 英語

    ポスター発表

  • 固定化したシトクロムP450発現大腸菌によるwhole cell bioconversion

    藤井 敦士, 石坂 明穂, 勝田 知尚, 今石 浩正, 山地 秀樹

    化学工学会第84年会, 2019年03月, 日本語, 化学工学会, 東京, 国内会議

    ポスター発表

  • Production of influenza virus-like particles in stably transformed insect cells

    MATSUDA Takuya, TANIJIMA Toshikazu, MASUMI-KOIZUMI Kyoko, KATSUDA Tomohisa, YAMAJI Hideki

    31st Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2018 Tsukuba), 2018年11月, 英語, 日本動物細胞工学会, 筑波, 国際会議

    ポスター発表

  • Production of antibody Fab fragment using 2A peptide in insect cells

    MIZOTE Yu, MASUMI-KOIZUMI Kyoko, KATSUDA Tomohisa, YAMAJI Hideki

    31st Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2018 Tsukuba), 2018年11月, 英語, 日本動物細胞工学会, 筑波, 国際会議

    ポスター発表

  • 緑藻Chlorella sorokinianaの光独立栄養培養における培養温度の影響

    桑井 仁葵, 山地 秀樹, 勝田 知尚

    化学工学会第50回秋季大会, 2018年09月, 日本語, 化学工学会, 鹿児島, 国内会議

    ポスター発表

  • 培養温度・光強度の周期的な変化が緑藻Haematococcus pluvialisの細胞分裂に及ぼす影響

    森本 高章, 山地 秀樹, 勝田 知尚

    化学工学会第50回秋季大会, 2018年09月, 日本語, 化学工学会, 鹿児島, 国内会議

    ポスター発表

  • 昆虫細胞における2Aペプチドを用いた抗体生産

    溝手 結, 増見 恭子, 勝田 知尚, 山地 秀樹

    第70回日本生物工学会大会, 2018年09月, 日本語, 日本生物工学会大会, 吹田, 国内会議

    口頭発表(一般)

  • ゲノム編集技術を用いた組換え昆虫細胞の作製

    戸上 真也, 増見 恭子, 勝田 知尚, 山地 秀樹

    化学工学会第50回秋季大会, 2018年09月, 英語, 化学工学会, 鹿児島, 国内会議

    ポスター発表

  • Production of influenza virus proteins in stably transformed insect cells

    YAMAJI Hideki, TANIJIMA Toshikazu, MATSUDA Takuya, Kyoko MASUMI-KOIZUMI, KATSUDA Tomohisa

    18th European Congress on Biotechnology (ECB2018), 2018年07月, 英語, European Federation of Biotechnology, Geneva, Switzerland, 国際会議

    ポスター発表

  • 緑藻Haematococcus pluvialisにおける周期的温度変化と細胞増殖の関係

    勝田 知尚, 森本 高章, 山地 秀樹

    化学工学会第83年会, 2018年03月, 日本語, 化学工学会, 吹田, 国内会議

    ポスター発表

  • 昆虫細胞によるインフルエンザウイルス抗原タンパク質の生産

    松田 拓也, 谷島 寿和, 増見 恭子, 勝田 知尚, 山地 秀樹

    化学工学会第83年会, 2018年03月, 日本語, 化学工学会, 吹田, 国内会議

    ポスター発表

  • 昆虫細胞による2Aペプチドを用いた抗体タンパク質の生産

    溝手 結, 増見 恭子, 勝田 知尚, 山地 秀樹

    化学工学会第49回秋季大会, 2017年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • 共焦点レーザー走査顕微鏡観察と数値ミュレーションに基づく抗体の吸着・破過の解析

    廣岡 奏, YAMAGUCHI Tomoki, 山地 秀樹, 勝田 知尚

    化学工学会第49回秋季大会, 2017年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • エアリフト式フォトバイオリアクターの操作条件と微細藻類の光独立栄養増殖の関係

    紅林 愛夏, 森本 高章, 西村 拓海, 山地 秀樹, 勝田 知尚

    化学工学会第49回秋季大会, 2017年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • Production of influenza virus proteins by recombinant insect cells

    YAMAJI Hideki, TANISHIMA Toshikazu, MASUMI-KOIZUMI Kyoko, KATSUDA Tomohisa

    International Meeting & 42nd Annual Conference of the Malaysian Society for Biochemistry & Molecular Biology (MSBMB), 2017年08月, 英語, The Malaysian Society for Biochemistry & Molecular Biology (MSBMB), Kuala Lumpur, Malaysia, 国際会議

    ポスター発表

  • 昆虫細胞を用いたインフルエンザウイルスタンパク質の生産

    山地 秀樹, 谷島 寿和, 増見 恭子, 勝田 知尚

    第30回日本動物細胞工学会2017年度大会 (JAACT2017), 2017年07月, 日本語, 日本動物細胞工学会, 松山, 国内会議

    ポスター発表

  • Efficient protein production by transient gene expression using insect cells

    YAMAJI Hideki, MORI Keita, HAMADA Hirotsugu, OHMURO Yuki, KATSUDA Tomohisa

    25th ESACT (the European Society for Animal Cell Technology) Meeting (ESACT 2017), 2017年05月, 英語, European Society for Animal Cell Technology (ESACT), Lausanne, Switzerland, 国際会議

    ポスター発表

  • リチウム添加による昆虫細胞からの抗体分泌生産の向上

    大室 有紀, 上田 宏, 山地 秀樹

    化学工学会第82年会, 2017年03月, 日本語, 化学工学会, 東京, 国内会議

    口頭発表(一般)

  • Transient gene expression in insect cells for efficient recombinant antibody production

    TAKAHASHI Ryoma, MASUMI-KOIZUMI Kyoto, OHMURO-MATSUYAMA Yuki, KATSUDA Tomohisa, YAMAJI Hideki

    The 29th Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2016 Kobe), 2016年11月, 英語, 日本動物細胞工学会, 神戸, 国際会議

    ポスター発表

  • Production of virus-like particles using recombinant insect cells

    TANISHIMA Toshikazu, KATSUDA Tomohisa, YAMAJI Hideki

    The 29th Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2016 Kobe), 2016年11月, 英語, 日本動物細胞工学会, 神戸, 国際会議

    ポスター発表

  • Lithium increases secretory production of antibody from recombinant insect cells

    OHMURO-MATSUYAMA Yuki, YAMAJI Hideki

    The 29th Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2016 Kobe), 2016年11月, 英語, 日本動物細胞工学会, 神戸, 国際会議

    口頭発表(一般)

  • Characterization of the diffusion and adsorption of IgG in protein A-coupled adsorbents with confocal laser scanning microscopy

    HIROOKA Kanamu, HARUNA Yusuke, YAMAJI Hideki, KATSUDA Tomohisa

    The 29th Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2016 Kobe), 2016年11月, 英語, 日本動物細胞工学会, 神戸, 国際会議

    ポスター発表

  • Taylor渦流れにおける細胞保持粒子の挙動と光合成微生物固定化の関係

    荷川取 将吾, 山地 秀樹, 大石 義彦, 河合 秀樹

    化学工学会第48回秋季大会, 2016年09月, 日本語, 化学工学会, 徳島, 国内会議

    口頭発表(一般)

  • Efficient antibody production by transient gene expression in insect cells

    YAMAJI Hideki, MORI Keita, HAMADA Hirotsugu, OHMURO Yuki, KATSUDA Tomohisa

    17th European Biotechnology Congress (ECB2016), 2016年07月, 英語, European Federation of Biotechnology (EFB), Krakow, Poland, 国際会議

    ポスター発表

  • Production of virus-like particles using insect cells

    YAMAJI Hideki

    The 2016 World Congress on In Vitro Biology, 2016年06月, 英語, Society for In Vitro Biology (SIVB), San Diego, USA, 国際会議

    [招待有り]

    口頭発表(招待・特別)

  • 培養条件の概日変化が緑藻Haematococcus pluvialisの栄養増殖に及ぼす影響

    吉峰 幸平, 山田 浩之, 勝田 知尚, 山地 秀樹

    化学工学会第81年会, 2016年03月, 日本語, 化学工学会, 大阪, 国内会議

    ポスター発表

  • 共焦点レーザー走査顕微鏡観察に基づくProtein A固定化吸着剤の拡散・吸着特性の評価

    勝田 知尚, 春名 祐佐, 廣岡 奏, 加瀬 裕貴, 大田 洸, 吉川 徹, 山地 秀樹

    化学工学会第81年会, 2016年03月, 日本語, 化学工学会, 大阪, 国内会議

    口頭発表(一般)

  • Recombinant antibody production by transient gene expression in insect cells

    YAMAJI Hideki, MORI Keita, HAMADA Hirotsugu, OHMURO Yuki, KATSUDA Tomohisa

    Asian Congress on Biotechnology 2015 (ACB2015), 2015年11月, 英語, Asian Federation of Biotechnolgy (AFOB), Kuala Lumpur, Malaysia, 国内会議

    ポスター発表

  • Photoautotrophic growth of Haematococcus pluvialis under controlled temperature and light conditions

    KATSUDA Tomohisa, GIANNELLI Luca, YAMADA Hiroyuki, YAMAJI Hideki

    Asian Congress on Biotechnology 2015 (ACB2015), 2015年11月, 英語, Asian Federation of Biotechnolgy (AFOB), Kuala Lumpur, Malaysia, 国内会議

    ポスター発表

  • Optimization of microalgal cultivation in cascade photobioreactors with wavy bottom via computational fluid dynamics

    WATANABE Chihiro, GIANNELLI Luca, YAMAJI Hideki, KATSUDA Tomohisa

    Asian Congress on Biotechnology 2015 (ACB2015), 2015年11月, 英語, Asian Federation of Biotechnolgy (AFOB), Kuala Lumpur, Malaysia, 国内会議

    ポスター発表

  • Modifications of endoplasmic reticulum signal sequence for antibody secretion from insect cells

    OHMURO Yuki, YAMAJI Hideki

    Asian Congress on Biotechnology 2015 (ACB2015), 2015年11月, 英語, Asian Federation of Biotechnolgy (AFOB), Kuala Lumpur, Malaysia, 国内会議

    ポスター発表

  • 昆虫細胞による抗体分泌生産のための小胞体シグナル配列の改変

    大室 有紀, 山地 秀樹

    第67回日本生物工学会大会, 2015年10月, 日本語, 日本生物工学会, 鹿児島, 国内会議

    ポスター発表

  • シトクロムP450発現大腸菌を用いたwhole cell bioconversionに及ぼす培養・反応条件の影響

    原田 真行, 森中 涼平, 勝田 知尚, 今石 浩正, 山地 秀樹

    化学工学会第47回秋季大会, 2015年09月, 日本語, 化学工学会, 札幌, 国内会議

    ポスター発表

  • Taylor渦バイオリアクターを目指した数値シミュレーションによる流動解析

    高橋 駿, 岡部 紫苑, 山地 秀樹, 河合 秀樹

    化学工学会第47回秋季大会, 2015年09月, 日本語, 化学工学会, 札幌, 国内会議

    口頭発表(一般)

  • 昆虫細胞を宿主とした一過性発現による抗体タンパク質の生産

    森 慶太, 濱田 宏嗣, 大室 有紀, 勝田 知尚, 山地 秀樹

    第28回日本動物細胞工学会2015年度大会 (JAACT2015), 2015年07月, 日本語, 日本動物細胞工学会, 仙台, 国内会議

    ポスター発表

  • 小胞体シグナル配列の改変が昆虫細胞による抗体分泌生産に及ぼす影響

    大室 有紀, 森 慶太, 濱田 宏嗣, 山地 秀樹

    第15回日本回蛋白質科学会年会, 2015年06月, 日本語, 日本回蛋白質科学会, 徳島, 国内会議

    ポスター発表

  • ヒカリコメツキムシ由来発光酵素Emerald lucを用いた蛋白質間相互作用検出系FlimPIAの構築

    下田 拓矢, 大室 有紀, 山地 秀樹, 上田 宏

    第15回日本回蛋白質科学会年会, 2015年06月, 日本語, 日本回蛋白質科学会, 徳島, 国内会議

    ポスター発表

  • Production of an antibody molecule by transient gene expression in insect cells

    MORI Keita, HAMADA Hirotsugu, OHMURO Yuki, KATSUDA Tomohisa, YAMAJI Hideki

    24th ESACT Meeting Barcelona 2015, 2015年06月, 英語, European Society for Animal Cell Technology (ESACT), Barcelona, Spain, 国際会議

    ポスター発表

  • カスケード型フォトバイオリアクターにおける微細藻類培養の数値流体力学に基づく検討

    渡邉 千尋, GIANNELLI Luca, 山地 秀樹, 勝田 知尚

    化学工学会姫路大会, 2014年12月, 日本語, 化学工学会関西支部, 姫路, 国内会議

    ポスター発表

  • 新規蛋白質間相互作用検出系FlimPIAの改良

    大室 有紀, 栗原 誠, 山下 貴宏, 綾部 圭一, 下田 拓矢, 山地 秀樹, 上田 宏

    生物発光化学発光研究会 第31回学術講演会, 2014年11月, 日本語, 生物発光化学発光研究会, 長野, 国内会議

    ポスター発表

  • バイオ医薬品開発と人材育成における神戸大の取り組み

    山地 秀樹

    第5回グライコバイオロジクス研究会, 2014年11月, 日本語, グライコバイオロジクス研究会, 神戸, 国内会議

    [招待有り]

    口頭発表(招待・特別)

  • Transient gene expression in insect cells for recombinant antibody production

    MORI Keita, HAMADA Hirotsugu, OGAWA Takahumi, OHMURO Yuki, KATSUDA Tomohisa, YAMAJI Hideki

    The 27th Annual Meeting of the Japanese Association for Animal Cell Technology (JAACT2014 Kitakyushu), 2014年11月, 英語, 日本動物細胞工学会, 北九州, 国際会議

    ポスター発表

  • Influence of substrate stiffness on pluripotency in mesenchymal cells

    EHASHI Tomo, TANIGUCHI Nami, YOSHIDA Kensuke, YAMAJI Hideki

    The 27th Annual Meeting of the Japanese Association for Animal Cell Technology (JAACT2014 Kitakyushu), 2014年11月, 英語, 日本動物細胞工学会, 北九州, 国際会議

    ポスター発表

  • 培養基材の弾性率が幹細胞の分化多能性に及ぼす影響

    江橋 具, 谷口 奈美, 吉田 健祐, 山地 秀樹

    化学工学会第46回秋季大会, 2014年09月, 日本語, 化学工学会, 福岡, 国内会議

    ポスター発表

  • 昆虫細胞を宿主とした一過性発現による抗体タンパク質生産

    濱田 宏嗣, 森 慶太, 小川 隆文, 大室 有紀, 勝田 知尚, 山地 秀樹

    化学工学会第46回秋季大会, 2014年09月, 日本語, 化学工学会, 福岡, 国内会議

    ポスター発表

  • 酵素改良と反応条件至適化による相互作用検出系FlimPIAの高機能化

    上田 宏, 栗原 誠, 山下 貴宏, 綾部 圭一, 山地 秀樹, 大室 有紀

    第66回日本生物工学会大会, 2014年09月, 日本語, 日本生物工学会, 札幌, 国内会議

    ポスター発表

  • Taylor渦バイオリアクターを目指した数値シミュレーションによる流動解析

    河合 秀樹, 赤沢 悠太, 山地 秀樹, 山田 雄大

    化学工学会第46回秋季大会, 2014年09月, 日本語, 化学工学会, 福岡, 国内会議

    口頭発表(一般)

  • Production of antibody fragments by transient gene expression in insect cells

    YAMAJI Hideki, KATSUDA Tomohisa, HAMADA Hirotsugu, MORI Keita

    16th European Congress on Biotechnology (ECB16), 2014年07月, 英語, European Federation of Biotechnology, Edinburgh, UK, 国際会議

    ポスター発表

  • 昆虫細胞を用いた一過性発現による機能性抗体タンパク質の生産

    森 慶太, 濵本 浩平, 宮口 尚子, 小川 隆文, 勝田 知尚, 山地 秀樹

    化学工学会第45回秋季大会, 2013年09月, 日本語, 化学工学会, 岡山市, 国内会議

    ポスター発表

  • シトクロムP450発現大腸菌の増殖と酵素活性に及ぼす培養条件の影響

    森中 涼平, 山地 秀樹, 今石 浩正

    第65回日本生物工学会大会, 2013年09月, 日本語, 日本生物工学会, 広島市, 国内会議

    ポスター発表

  • Taylor-Couette渦バイオリアクターにおける光合成微生物の増殖特性

    岡部 紫苑, 河合 秀樹, 山地 秀樹, 中村 恵龍

    化学工学会第45回秋季大会, 2013年09月, 日本語, 化学工学会, 岡山市, 国内会議

    ポスター発表

  • Haematococcus pluvialisの増殖とアスタキサンチン生産に及ぼす培養温度の影響

    山田 浩之, 和田 壮太郎, GIANNELLI Luca, 山地 秀樹, 勝田 知尚

    化学工学会第45回秋季大会, 2013年09月, 日本語, 化学工学会, 岡山市, 国内会議

    ポスター発表

  • Efficient production of viral envelope proteins in recombinant insect cells

    YAMAJI Hideki, KUWAHARA Miwa, KONISHI Eiji

    2013 ESACT Meeting in Lille, 2013年06月, 英語, European Society for Animal Cell Technology (ESACT), Lille, France, 国際会議

    ポスター発表

  • Photobioreactor design using CFD: mixing and light effects on microalgal cultures

    GIANNELLI Luca, WADA Sotaro, YAMADA Hiroyuki, YAMAJI Hideki, KATSUDA Tomohisa

    化学工学会第78年会, 2013年03月, 英語, 化学工学会, 豊中市, 国内会議

    口頭発表(一般)

  • Production of recombinant antibody molecules in insect cells

    MIYAGUCHI Naoko, HAMAMOTO Kouhei, OGAWA Takafumi, SONODA Hiroyuki, KATSUDA Tomohisa, YAMAJI Hideki

    The 25th Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2012), 2012年11月, 英語, 日本動物細胞工学会, 名古屋市, 国際会議

    ポスター発表

  • Production and purification of Japanese encephalitis virus-like particles from recombinant insect cells

    HOTTA Yuri, MINAMITANI Azusa, NARITA Kazuma, KATSUDA Tomohisa, YAMAJI Hideki

    The 25th Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2012), 2012年11月, 英語, 日本動物細胞工学会, 名古屋市, 国際会議

    ポスター発表

  • 組換え昆虫細胞によるscFv-Fc融合タンパク質の生産

    薗田 啓之, 熊田 陽一, 勝田 知尚, 山地 秀樹

    第64回日本生物工学会大会, 2012年10月, 日本語, 日本生物工学会, 神戸市, 国内会議

    口頭発表(一般)

  • Computational fluid dynamics for enhancing the light distribution in a photobioreactor grown culture of Hematococcus pluvialis

    GIANNELLI Luca, WADA Sotaro, YAMAJI Hideki, KATSUDA Tomohisa

    第64回日本生物工学会大会, 2012年10月, 英語, 日本生物工学会, 神戸市, 国内会議

    口頭発表(一般)

  • 大腸菌を用いた一本鎖抗体生産における終止コドンの影響

    山條 佑樹, 福丸 由夏, 山地 秀樹, 勝田 知尚

    化学工学会第44回秋季大会, 2012年09月, 日本語, 化学工学会, 仙台市, 国内会議

    ポスター発表

  • Taylor-Couette渦バイオリアクターにおける光合成微生物の増殖と藻体破壊について

    小林 裕平, 河合 秀樹, 山地 秀樹, 中村 恵龍

    化学工学会第44回秋季大会, 2012年09月, 日本語, 化学工学会, 仙台市, 国内会議

    ポスター発表

  • Secretory production of virus-like particles by recombinant insect cells

    YAMAJI Hideki, KUWAHARA Miwa, KONISHI Eiji

    The 15th European Congress on Biotechnology of the European Federation of Biotechnology (ECB15), 2012年09月, 英語, The European Federation of Biotechnology, Istanbul, Turkey, 国際会議

    ポスター発表

  • 昆虫細胞-バキュロウイルス系によるウイルス様粒子の生産

    山地 秀樹, 瀬川 麻衣子, 中村 匡崇, 勝田 知尚

    化学工学会第77年会, 2012年03月, 日本語, 化学工学会, 東京, 国内会議

    口頭発表(一般)

  • 炭化水素資化酵母Yarrowia lipolyticaによる油汚染土壌の修復のための無機栄養源の開発

    藤本 翔, 田沼 直樹, 勝田 知尚, 山地 秀樹

    化学工学会第43回秋季大会, 2011年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • 昆虫細胞を用いた抗体タンパク質の分泌生産

    濵本 浩平, 宮口 尚子, 澤本 詩織, 小川 隆文, 勝田 知尚, 山地 秀樹

    化学工学会第43回秋季大会, 2011年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • 昆虫細胞によるウイルス様粒子生産に及ぼす培養条件の検討

    成田 和馬, 堀田 有里, 南谷 梓, 永菅 尚, 勝田 知尚, 山地 秀樹

    化学工学会第43回秋季大会, 2011年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • シャイン・ダルガーノ配列を改変した大腸菌による一本鎖抗体の流加培養生産

    福丸 由夏, 野上 達弘, 木原 麻那, 勝田 知尚, 山地 秀樹

    化学工学会第43回秋季大会, 2011年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • Taylor-Couette渦バイオリアクターにおける光合成微生物の固定化の効果について

    河合 秀樹, 中村 恵龍, 山地 秀樹

    化学工学会第43回秋季大会, 2011年09月, 日本語, 化学工学会, 名古屋, 国内会議

    ポスター発表

  • Evaluation of extracellular subviral particles of dengue type 2 virus produced by insect cells for use as vaccine and diagnostic antigens

    KUWAHARA Miwa, YAMAJI Hideki, KONISHI Eiji

    IUMS 2011 Sapporo Congress, 2011年09月, 英語, International Union of Microbiological Societies, 札幌, 国際会議

    ポスター発表

  • Efficient production of extracellular subviral particles of Japanese encephalitis virus by recombinant insect cells

    YAMAJI Hideki, NAGASUGA Takashi, TAKAHASHI Yusuke, NAKAMURA Masataka, KATSUDA Tomohisa, KUWAHARA Miwa, KONISHI Eiji

    IUMS 2011 Sapporo Congress, 2011年09月, 英語, International Union of Microbiological Societies, 札幌, 国内会議

    ポスター発表

  • Secretory production of virus-like particles by recombinant insect cells

    YAMAJI Hideki, NAGASUGA Takashi, TAKAHASHI Yusuke, NAKAMURA Masataka, KATSUDA Tomohisa, KUWAHARA Miwa, KONISHI Eiji

    ESACT 2011 Meeting, 2011年05月, 英語, European Society for Animal Cell Technology, Vienna, Austria, 国際会議

    ポスター発表

  • 炭化水素資化酵母Yarrowia lipolyticaを用いた油汚染土壌の修復における栄養源の添加条件の検討

    藤本 翔, 田沼 直樹, 勝田 知尚, 山地 秀樹

    第13回化学工学会学生発表会(神戸大会), 2011年03月, 日本語, 化学工学会, 神戸市, 国内会議

    口頭発表(一般)

  • 大腸菌を用いた一本鎖抗体生産におけるシャイン・ダルガーノ配列の影響

    福丸 由夏, 野上 達弘, 勝田 知尚, 山地 秀樹

    第13回化学工学会学生発表会(神戸大会), 2011年03月, 日本語, 化学工学会, 神戸市, 国内会議

    口頭発表(一般)

  • 大腸菌におけるインクルージョンボディ生成に及ぼすシャイン・ダルガーノ配列の影響

    勝田 知尚, 福丸 由夏, 木原 麻那, 野上 達弘, 山地 秀樹

    化学工学会第76年会, 2011年03月, 日本語, 化学工学会, 東京都, 国内会議

    口頭発表(一般)

  • Optimized Shine-Dalgarno sequence for efficient production of single-chain Fv antibody in Escherichia coli

    NOGAMI Tatsuhiro, KIHARA Mana, SANO Tadahisa, KATSUDA Tomohisa, YAMAJI Hideki

    The International Chemical Congress of Pacific Basin Societies (Pacifichem 2010), 2010年12月, 英語, Canadian Society for Chemistry, American Chemical Society, Chemical Society of Japan, New Zealand Institute of Chemistry, Royal Australian Chemical Institute, Korean Chemical Society, Chinese Chemical Society, Honolulu, USA, 国際会議

    ポスター発表

  • Rapid production of functional antibody molecules by transient gene expression in insect cells

    SAWAMOTO Shiori, HAMAMOTO Kouhei, KATSUDA Tomohisa, YAMAJI Hideki

    Young Asian Biochemical Engineers Community (YABEC) Symposium 2010, 2010年11月, 英語, Young Asian Biochemical Engineers Community (YABEC), Taipei, Taiwan, 国際会議

    ポスター発表

  • Effect of signal peptide on the production of single-chain fv antibody by Escherichia coli

    NOGAMI Tatsuhiro, FUKUMARU Yuka, KATSUDA Tomohisa, YAMAJI Hideki

    Young Asian Biochemical Engineers Community (YABEC) Symposium 2010, 2010年11月, 英語, Young Asian Biochemical Engineers Community (YABEC), Taipei, Taiwan, 国際会議

    ポスター発表

  • Production of Japanese encephalitis virus-like particles in insect cell expression systems

    山地 秀樹, 永菅 尚, 高橋 裕輔, 中村 匡崇, 勝田 知尚, 桑原 三和, 小西 英二

    The 13th Asia Pacific Confederation of Chemical Engineering Congress (APCChE 2010), 2010年10月, 英語, Asia Pacific Confederation of Chemical Engineering, Taipei, Taiwan, 国際会議

    ポスター発表

  • 培養昆虫細胞を用いた有用物質生産

    山地 秀樹

    化学工学会第42回秋季大会, 2010年09月, 日本語, 化学工学会, 京都市, 国内会議

    その他

  • プロテインAアフィニティクロマトグラフィーのための溶離条件検討法の開発

    加瀬 裕貴, 西山 拓也, 勝田 知尚, 山地 秀樹

    化学工学会第42回秋季大会, 2010年09月, 日本語, 化学工学会, 京都市, 国内会議

    ポスター発表

  • シャペロン共発現による大腸菌での一本鎖抗体 (scFv) 融合タンパク質の生産

    薗田 啓之, 熊田 陽一, 勝田 知尚, 山地 秀樹

    化学工学会第42回秋季大会, 2010年09月, 日本語, 化学工学会, 京都市, 国内会議

    ポスター発表

  • アルカン資化酵母Yarrowia lipolyticaを用いた土壌汚染油の分解

    田沼 直樹, 藤本 翔, 塩見 尚史, 勝田 知尚, 山地 秀樹

    化学工学会第42回秋季大会, 2010年09月, 日本語, 化学工学会, 京都市, 国内会議

    ポスター発表

  • Production of Japanese encephalitis virus-like particles using insect cells

    永菅 尚, 高橋 裕輔, 中村 匡崇, 勝田 知尚, 山地 秀樹

    The 23rd Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2010), 2010年09月, 英語, The Japanese Association for Animal Cell Technology, 札幌市, 国際会議

    ポスター発表

  • Pichia pastorisを用いた一本鎖抗体生産における誘導初期のメタノール濃度の影響

    藤田 啓介, 秋山 直毅, 勝田 知尚, 山地 秀樹

    化学工学会第75年会, 2010年03月, 日本語, 化学工学会, 鹿児島, 国内会議

    口頭発表(一般)

  • 昆虫細胞-バキュロウイルス系を用いた抗体タンパク質の高効率生産

    澤本 詩織, 古田 貴憲, 勝田 知尚, 山地 秀樹

    第8回最先端バイオテクノロジー発表会, 2010年02月, 日本語, 化学工学会, 大阪, 国内会議

    ポスター発表

  • 一本鎖抗体の可溶性発現に及ぼすSD配列改変の影響

    野上 達弘, 木原 麻那, 佐野 宰久, 勝田 知尚, 山地 秀樹

    第8回最先端バイオテクノロジー発表会, 2010年02月, 日本語, 化学工学会, 大阪, 国内会議

    ポスター発表

  • Effects of Shine-Dalgarno sequence on the production of single-chain Fv antibody by Escherichia coli

    KIHARA Mana, NOGAMI Tatsuhiro, SANO Tadahisa, KATSUDA Tomohisa, YAMAJI Hideki

    Young Asian Biochemical Engineers Community (YABEC) Symposium 2009, 2009年12月, 英語, Young Asian Biochemical Engineers Community, Xiamen, 国際会議

    ポスター発表

  • Production of Japanese encephalitis virus-like particles by recombinant insect cells

    山地 秀樹, 高橋 裕輔, 中村 匡崇, 勝田 知尚, 桑原 三和, 小西 英二

    Asia Pacific Biochemical Engineering Conference 2009 (APBioChEC '09), 2009年11月, 英語, APBioChEC '09 Committee, 神戸市, 国際会議

    口頭発表(一般)

  • Oil degradation in soil by a hydrocarbon assimilating yeast, Yarrowia lipolytica

    FUKUI Mayu, SHIOMI Shinji, TANUMA Naoki, KATSUDA Tomohisa, SHIOMI Naofumi, YAMAJI Hideki

    The 9th Asia-Pacific Biochemical Engineering Conference 2009, 2009年11月, 英語, The 9th Asia-Pacific Biochemical Engineering Conference 2009, Kobe, 国際会議

    ポスター発表

  • 大腸菌を用いた一本鎖抗体生産におけるSD配列の影響

    野上 達弘, 木原 麻那, 佐野 宰久, 勝田 知尚, 山地 秀樹

    化学工学会第41回秋季大会, 2009年09月, 日本語, 化学工学会, 東広島市, 国内会議

    ポスター発表

  • 昆虫細胞を用いた日本脳炎ウイルス様粒子の生産

    高橋 裕輔, 永菅 尚, 中村 匡崇, 勝田 知尚, 山地 秀樹

    第61回日本生物工学会大会, 2009年09月, 日本語, 日本生物工学会, 名古屋市, 国内会議

    口頭発表(一般)

  • 昆虫細胞-バキュロウイルス系による抗体タンパク質の生産

    澤本 詩織, 鈴木 輔, 古田 貴憲, 小川 隆文, 勝田 知尚, 山地 秀樹

    化学工学会第41回秋季大会, 2009年09月, 日本語, 化学工学会, 東広島市, 国内会議

    ポスター発表

  • ヘマトコッカス藻の高密度培養に対する二酸化チタン-超音波励起法の適用

    岩崎 大記, 千賀 至, 山下 貴史, 東内 雅博, 荻野 千秋, 清水 宣明, 勝田 知尚, 山地 秀樹

    化学工学会第41回秋季大会, 2009年09月, 日本語, 化学工学会, 東広島市, 国内会議

    ポスター発表

  • 昆虫細胞を用いた日本脳炎ウイルスタンパク質の生産

    高橋 裕輔, 中村 匡崇, 古田 貴憲, 勝田 知尚, 山地 秀樹

    化学工学会第74年会, 2009年03月, 日本語, 化学工学会, 横浜市, 国内会議

    口頭発表(一般)

  • 一塩基変異を検出する昇温溶出アフィニティクロマトグラフィーに対するPCRの適用性

    勝田 知尚, 今西 誠, 加藤 滋雄, 山地 秀樹

    化学工学会第74年会, 2009年03月, 日本語, 化学工学会, 横浜市, 国内会議

    口頭発表(一般)

  • Supplementation of phosphatidylcholine protects the hybridoma cells from apoptosis induced by serum withdrawal

    TEY Beng Ti, YAP Kah Choon, YAMAJI Hideki, ALI A. Manaf, TAN Wen Siang

    The 21th Annual and International Meeting of the Japanese Association for Animal Cell Technology, 2008年11月, 英語, 日本動物細胞工学会, 福岡市, 国際会議

    ポスター発表

  • Production of adenovirus vector by 293 cells immobilized within porous biomass support particles

    MORISHITA Naoya, KUBO Shuji, GOTOH Akinobu, KATSUDA Tomohisa, FUKUDA Hideki, YAMAJI Hideki

    The 21th Annual and International Meeting of the Japanese Association for Animal Cell Technology, 2008年11月, 英語, 日本動物細胞工学会, 福岡市, 国際会議

    ポスター発表

  • Low multiplicity infection of 293 cells for adenovirus vector production

    MORISHITA Naoya, YAMADA Kentaro, KUBO Shuji, GOTOH Akinobu, KATSUDA Tomohisa, FUKUDA Hideki, YAMAJI Hideki

    The 21th Annual and International Meeting of the Japanese Association for Animal Cell Technology, 2008年11月, 英語, 日本動物細胞工学会, 福岡市, 国際会議

    ポスター発表

  • High efficiency expression of soluble scFv in E. coli by use of linkers containing rare codons

    KAJIHARA Hideyuki, SAKAN Yoshinobu, KIHARA Mana, KIKUCHI Yasufumi, KUMADA Yoichi, KATSUDA Tomohisa, YAMAJI Hideki, KATOH Shigeo

    The 14th Symposium of Young Asian Biochemical Engineers’ Community, 2008年11月, 英語, Young Asian Biochemical Engineers’ Community, 東京都, 国際会議

    ポスター発表

  • Secretory production of Japanese encephalitis virus E protein by recombinant insect cells

    TAKAHASHI Yusuke, NAKAMURA Masataka, FURUTA Takanori, KATSUDA Tomohisa, YAMAJI Hideki

    “Bioseparation for Biorecognition and Bionanotechnology” Conference, 2008年10月, 英語, Hanyang University, Ansan, Korea, 国内会議

    ポスター発表

  • Detection of single nucleotide variation in amplified gene fragments using oligonucleotide-coupled affinity column

    KATSUDA Tomohisa, IMANISHI Makoto, YAMAJI Hideki, KATOH Shigeo

    “Bioseparation for Biorecognition and Bionanotechnology” Conference, 2008年10月, 英語, Hanyang University, Ansan, Korea, 国内会議

    ポスター発表

  • 多孔性粒子に固定化した大腸菌を用いる有用物質生産

    金 娜, 勝田 知尚, 福田 秀樹, 山地 秀樹

    化学工学会第40回秋季大会, 2008年09月, 日本語, 化学工学会, 仙台市, 国内会議

    ポスター発表

  • レアコドンリンカーによる大腸菌での可溶性scFvの高効率生産

    木原 麻那, 佐官 義信, 梶原 英之, 赤坂 翔, 勝田 知尚, 加藤 滋雄, 山地 秀樹

    化学工学会第40回秋季大会, 2008年09月, 日本語, 化学工学会, 仙台市, 国内会議

    ポスター発表

  • 293細胞の固定化培養によるアデノウイルスベクターの生産

    森下 直矢, 山田 健太郎, 山地 秀樹, 福田 秀樹

    化学工学会第73年会, 2008年03月, 日本語, 化学工学会, 浜松, 国内会議

    口頭発表(一般)

  • リンカーの改変による一本鎖抗体可溶成分の効率的生産

    佐官 義信, 川﨑 朋美, 梶原 英之, 山地 秀樹, 菊池 康文, 熊田 陽一, 加藤 滋雄

    化学工学会山口大会, 2007年11月, 日本語, 化学工学会中国四国支部, 宇部, 国内会議

    ポスター発表

  • High efficiency expression of soluble scFv in E. coli by use of designed linkers

    SAKAN Yoshinobu, KAWASAKI Tomomi, KAJIHARA Hideyuki, YAMAJI Hideki, KIKUCHI Yasuhumi, KUMADA Yoichi, KATOH Shigeo

    Asia Pacific Biochemical Engineering Conference 2007, 2007年11月, 英語, Biochemical Engineering Society of Taiwan, Taipei, Taiwan, 国際会議

    ポスター発表

  • 昆虫細胞を用いる機能性抗体の生産システム

    古田 貴憲, 山地 秀樹, 福田 秀樹

    化学工学会第39回秋季大会, 2007年09月, 日本語, 化学工学会, 札幌, 国内会議

    ポスター発表

  • Display of functional antibody Fab fragment on the baculovirus surface

    YAMAJI Hideki, FUKUDA Hideki

    13th European Congress on Biotechnology, 2007年09月, 英語, European Federation of Biotechnology, Barcelona, Spain, 国際会議

    ポスター発表

  • 機能性タンパク質を表面に提示したバキュロウイルスの作製

    高田 新也, 山地 秀樹, 福田 秀樹

    化学工学会第72年会, 2007年03月, 日本語, 化学工学会, 京都大学, 国内会議

    口頭発表(一般)

  • 多孔性粒子を用いる大腸菌の固定化培養

    黄 娟, 山地 秀樹, 福田 秀樹

    化学工学会第38回秋季大会, 2006年09月, 日本語, 化学工学会, 福岡大学, 国内会議

    口頭発表(一般)

  • Whole cell biocatalystによるバイオディーゼル燃料生産

    沼田 崇男, 福水 崇裕, 濵 真司, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    化学工学会第38回秋季大会, 2006年09月, 日本語, 化学工学会, 福岡大学, 国内会議

    口頭発表(一般)

  • Production of functional antibody Fab fragment in stably transformed insect cells

    YAMAJI Hideki, FUKUDA Hideki

    JAACT 2006 Kyoto (The 19th Annual and International Meeting of the Japanese Association for Animal Cell Technology), 2006年09月, 英語, 日本動物細胞工学会, 京都, 国際会議

    ポスター発表

  • 昆虫細胞培養による有用タンパク質生産

    山地 秀樹, 福田 秀樹

    化学工学会室蘭大会, 2006年08月, 日本語, 化学工学会, 室蘭工業大学, 国内会議

    口頭発表(一般)

  • Production of functional antibody molecules in recombinant insect cells

    YAMAJI Hideki, FUKUDA Hideki

    Antibody Engineering Conference, 2006年06月, 英語, 抗体エンジニアリング国際会議事務局, 第20回国際生化学・分子生物学会議, 鹿児島, 国際会議

    ポスター発表

  • 微生物充填カラムバイオリアクターを用いたバイオディーゼル燃料生産

    福水 崇裕, 濱 真司, 沼田 崇男, 野田 秀夫, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    化学工学会第71年会, 2006年03月, 日本語, 化学工学会, 東京工業大学, 国内会議

    口頭発表(一般)

  • Whole cell biocatalystsによるリン脂質sn-1部位のエステル交換反応

    三浦 主寛, 濱 真司, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    化学工学会第71年会, 2006年03月, 日本語, 化学工学会, 東京工業大学, 国内会議

    口頭発表(一般)

  • Aspergillus oryzaeのタンパク質分泌プロセスにおけるROL由来N末端アミノ酸配列の役割

    濱 真司, 進戸 尚樹, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    化学工学会第71年会, 2006年03月, 日本語, 化学工学会, 東京工業大学, 国内会議

    口頭発表(一般)

  • 組換え昆虫細胞を用いた抗体Fabフラグメントの分泌生産

    山地 秀樹, 眞鍋 利孝, 福田 秀樹

    平成17年度日本生物工学会大会, 2005年11月, 日本語, 化学工学会, つくば国際会議場, 国内会議

    口頭発表(一般)

  • 微生物充填式カラムバイオリアクターを用いたバイオディーゼル燃料生産

    沼田 崇男, 福水 崇裕, 濱 真司, 進戸 尚樹, 野田 秀夫, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    第8回化学工学会学生発表会, 2005年09月, 日本語, 化学工学会, 広島大学, 国内会議

    ポスター発表

  • 多孔性粒子を用いた大腸菌の固定化

    黄 娟, 山地 秀樹, 福田 秀樹

    化学工学会第37回秋季大会, 2005年09月, 日本語, 化学工学会, 岡山大学, 国内会議

    口頭発表(一般)

  • Aspergillus oryzaeにおけるリパーゼ分泌プロセスの可視化

    進戸 尚樹, 濱 真司, 沼田 崇男, 山地 秀樹, 近藤 昭彦, 福田 秀樹

    第8回化学工学会学生発表会, 2005年09月, 日本語, 化学工学会, 広島大学, 国内会議

    ポスター発表

  • 昆虫細胞を用いた有用タンパク質の高生産プロセス

    山地 秀樹

    日本動物細胞工学会2005年度大会, 2005年07月, 日本語, 日本動物細胞工学会, 東京大学, 国内会議

    口頭発表(招待・特別)

  • Immobilized cell culture using different media for efficient production of biochemicals

    YAMAJI Hideki, NODA Hideo, FUKUDA Hideki

    7th World Congress of Chemical Engineering, 2005年07月, 英語, Institution of Chemical Engineers, グラスゴー, 国際会議

    口頭発表(一般)

  • 組換え昆虫細胞を用いた抗体Fabフラグメントの分泌発現

    真鍋 利孝, 山地 秀樹, 藤井 郁雄, 福田 秀樹

    化学工学会 第70年会, 2005年, 日本語, 化学工学会, 名古屋大学, 国内会議

    口頭発表(一般)

  • 昆虫細胞-バキュロウイルス系による抗体Fabフラグメントの生産

    阪本 真, 真鍋 利孝, 山地 秀樹, 福田 秀樹

    第7回化学工学会学生発表会, 2005年, 日本語, 化学工学会, 同志社大学, 国内会議

    口頭発表(一般)

  • 組換え昆虫細胞を用いた抗体Fabフラグメントの生産

    真鍋 利孝, 山地 秀樹, 藤井 郁雄, 福田 秀樹

    日本生物工学会大会, 2004年09月, 日本語, 日本生物工学会, 名城大学, 国内会議

    口頭発表(一般)

  • Efficient recombinant protein production in insect cell culture

    YAMAJI Hideki, FUKUDA Hideki

    YABEC 2004, 2004年09月, 英語, 未記入, 大阪, 国際会議

    口頭発表(一般)

  • 昆虫細胞を用いた抗体Fabフラグメントの生産

    真鍋 利孝, 渡壁 慶三, 山地 秀樹, 藤井 郁雄, 福田 秀樹

    化学工学会関西支部第2回最先端バイオテクノロジー公開セミナー, 2004年, 日本語, 化学工学会関西支部, 神戸女学院大学, 国内会議

    口頭発表(一般)

  • 昆虫細胞のバイオリアクター技術

    山地 秀樹

    化学工学会環境部会リサイクル分科会主催若手研究者による資源リサイクルに関する研究発表会, 2004年, 日本語, 化学工学会環境部会, 未記入, 国内会議

    口頭発表(一般)

  • 固定化昆虫細胞による組換えタンパク質の生産

    真鍋 利孝, 北浦 明法, 山地 秀樹, 福田 秀樹

    化学工学会第69年会, 2004年, 日本語, 化学工学会, 大阪府立大学, 国内会議

    口頭発表(一般)

  • Efficient protein production by insect cells immobilized within porous support particles

    MANABE Toshitaka, KITAURA Akinori, YAMAJI Hideki, FUKUDA Hideki

    The 10th APCChE Congress, 2004年, 英語, 未記入, 北九州, 国際会議

    口頭発表(一般)

  • Efficient Production from Cellulosic Materials by Cell Surface Engineered Yeast Strains

    MANABE Toshitaka, KITAURA Akinori, YAMAJI Hideki, FUKUDA Hideki

    The 10th APCChE Congress, 2004年, 英語, 不明, 不明, 国際会議

    口頭発表(一般)

  • 血清無添加の基礎培地における昆虫細胞ーバキュロウイルス系によるタンパク質生産

    西川 典克, 山地 秀樹, 福田 秀樹

    化学工学会第68年会, 2003年03月, 日本語, 化学工学会, 東京, 国内会議

    口頭発表(一般)

  • 無血清培養下におけるCHO細胞に及ぼすホスファチジン酸の効果

    川淵 智子, 城宝 司, 林 智佐, 坂井 健太郎, 山地 秀樹, 福田 秀樹

    日本動物細胞工学会2003年大会, 2003年, 日本語, 日本動物細胞工学会, 大阪, 国内会議

    口頭発表(一般)

  • 動物・昆虫細胞培養による有用タンパク質生産の効率化

    山地 秀樹, 福田 秀樹

    化学工学会京都大会, 2003年, 日本語, 化学工学会, 京都, 国内会議

    口頭発表(一般)

  • 昆虫細胞による組換えタンパク質生産に及ぼす血清無添加の基礎培地への培地交換の影響

    北浦 明法, 西川 典克, 山地 秀樹, 福田 秀樹

    平成15年度日本生物工学会大会, 2003年, 日本語, 日本生物工学会, 熊本大学, 国内会議

    口頭発表(一般)

  • Recombinant protein production by insect cells immobilized within porous support particles in basal synthetic medium

    YAMAJI Hideki, FUKUDA Hideki

    11th European Congress on Biotechnology, 2003年, 英語, 未記入, 未記入, 国際会議

    口頭発表(一般)

  • 遺伝子組換え昆虫細胞樹立へのゲノム編集技術の応用

    松田 拓也, 戸上 真也, 増見 恭子, Prihardi Kahar, 勝田 知尚, 山地 秀樹

    化学工学会第52回秋季大会

  • ゲノム編集技術を用いた遺伝子組換え昆虫細胞の樹立

    松田 拓也, 戸上 真也, 増見 恭子, Prihardi Kahar, 勝田 知尚, 山地 秀樹

    第34回日本動物細胞工学会2021年度大会 (JAACT2021)

  • プロテインA固定化マイクロアフィニティカラムを用いた疑似オンラインIgG濃度測定法の開発

    川村 和樹, 勝田 知尚, 山地 秀樹

    化学工学会第86年会

  • Production of influenza virus-like particles using recombinant insect cells

    Takuya Matsuda, Toshikazu Tanijima, Akito Hirose, Kyoko Masumi-Koizumi, Tomohisa Katsuda, Hideki Yamaji

    The 33rd Annual and International Meeting of the Japanese Association for Animal Cell Technology (JAACT2020 Fuchu)

  • 昆虫細胞を用いたインフルエンザウイルス様粒子の生産

    松田 拓也, 増見 恭子, 勝田 知尚, 山地 秀樹

    化学工学会第51回秋季大会

  • 異種タンパク質を提示するインフルエンザウイルス様粒子の昆虫細胞を用いた生産

    市川裕大, 松田拓也, Prihardi Kahar, 勝田知尚, 山地秀樹

    化学工学会第87年会, 2022年03月, 日本語

    ポスター発表

  • プロテインA担体における抗体の拡散・吸着挙動の検討

    林航平, 勝田知尚, 山地秀樹

    化学工学会第87年会, 2022年03月, 日本語

    ポスター発表

  • 光合成微生物の生育に及ぼす環境条件の影響

    河合秀樹, 大石義彦, 今野匠, 山地秀樹

    第31回化学工学・粉体工学研究発表会, 2022年01月, 日本語

    口頭発表(一般)

共同研究・競争的資金等の研究課題

  • 組換え昆虫細胞によるアデノ随伴ウイルスベクター生産のための最適分子設計

    山地 秀樹

    日本学術振興会, 科学研究費助成事業 基盤研究(B), 基盤研究(B), 神戸大学, 2019年04月 - 2022年03月

  • 山地 秀樹

    科学研究費補助金/基盤研究(B), 2015年04月 - 2019年03月, 研究代表者

    競争的資金

  • 山地 秀樹

    科学研究費補助金/萌芽研究, 2012年04月 - 2014年03月, 研究代表者

    競争的資金

  • 山地 秀樹

    科学研究費補助金/基盤研究(B), 2010年, 研究代表者

    競争的資金

  • 山地 秀樹

    科学研究費補助金/基盤研究(C), 2007年, 研究代表者

    競争的資金

  • 山地 秀樹

    科学研究費補助金/基盤研究(C), 2005年, 研究代表者

    競争的資金

  • 福田 秀樹, 山地 秀樹, 後藤章亮暢, 白川 利朗

    科学研究費補助金/萌芽研究, 2005年

    競争的資金

  • プロテオーム解析による新規細胞培養技術の開発

    山地 秀樹, 福田 秀樹

    日本学術振興会, 科学研究費助成事業 基盤研究(C), 基盤研究(C), 神戸大学, 2002年 - 2003年

    動物細胞や昆虫細胞の培養による有用タンパク質の高生産プロセスを構築するために最も重要な技術課題は,培養の無血清化である.本研究では,培養の無血清化にともなう細胞のプロテオームの変化を解析することにより,無血清化に関与するタンパク質の同定と発現メカニズムの解明を行い,無血清化のプロセスを迅速化,効率化することを最終的な目標として,基礎的なデータの収集を行った.まず,有用物質生産のための遺伝子組換えの宿主細胞として広く用いられている動物細胞であるCHO細胞や昆虫細胞Sf9の低コストの無血清培養技術を開発するために,細胞増殖や組換えタンパク質生産を促進する効果を有するリン脂質であるホスファチジン酸の供給方法や血清無添加の基礎培地における昆虫細胞-バキュロウイルス系による組換えタンパク質生産などについて検討を行った.また,CHO細胞については,細胞を継代する際に使用する培地の血清濃度を徐々に低下させることによって血清添加培地を無血清培地に代替していく馴化法により,無血清培地中で生育可能な細胞を取得した.得られた血清非依存性細胞の増殖速度や組換えタンパク質生産速度を測定し,血清添加培地を用いて継代している細胞のそれらとほぼ同等であることを確認した.次に,無血清培地への馴化にともないCHO細胞のプロテオームがどのように変化したのかを調べるために,血清非依存性細胞および血清添加培地を用いて継代している細胞の細胞内タンパク質を2次元電気泳動により解析した.その結果,計約700種類の細胞内タンパク質が検出され,培養の無血清化にともない発現量が5倍に増加または1/5に減少したタンパク質がそれぞれ数種類ずつ存在することがわかった.

  • 昆虫細胞培養による情報伝達関連タンパク質生産の特性解析と効率化

    山地 秀樹

    日本学術振興会, 科学研究費助成事業 奨励研究(A), 奨励研究(A), 神戸大学, 2000年 - 2001年

    昆虫細胞-バキュロウイルス系は,生物学的活性や高次構造が本来のものと同様の組換えタンパク質を大量に発現可能な系として近年注目を集めている.昆虫細胞の培養では,動物細胞の場合と同様に,通常,牛胎児血清(FBS)を10%程度添加した培地が用いられる.しかしながら,血清の使用により,目的タンパク質の分離精製が困難となるなどの問題が生じるため,工業的規模での有用タンパク質の生産には無血清培地の使用が望まれる.昆虫細胞培養による機能性タンパク質の高効率生産プロセスを構築するための基礎を確立することを目的として,本年度は,血清を添加していない基礎培地における組換えタンパク質生産について検討した. まず,基礎培地としてTNM-FHを用いてFBSの有無が昆虫細胞Sf9の増殖に及ぼす影響を調べた.FBSを10%添加したTNM-FHと比べて,FBSを添加していない場合,昆虫細胞Sf9の増殖速度は小さく,到達細胞密度も低かった.また,動物細胞の増殖促進効果を有するホスファチジン酸(PA)を添加しても,細胞増殖には変化が見られなかった. ところが,β-galactosidaseを発現する組換え核多角体病ウイルスをSf9に感染させ組換えタンパク質生産に及ぼす影響を検討したところ,FBSを添加していないTNM-FHの場合でも,FBSを10%添加した場合と比較して約2/3のβ-galactosidaseが生産されることがわかった。また,PAを10mg/lの濃度で添加する,あるいは組換えウイルスの感染多重度(MOI)を高くすることにより,いずれもβ-galactosidaseの生産がさらに促進された.このように,血清を添加していない基礎培地を用いた場合でも,大量の組換えタンパク質を生産できたことから,経済性などの点で優れたプロセスを構築できる可能性が示唆された.

  • 固定化昆虫細胞によるタンパク質の高生産バイオリアクタープロセスの開発

    福田 秀樹, 橘 佳永, 山地 秀樹

    日本学術振興会, 科学研究費助成事業 基盤研究(B), 基盤研究(B), 神戸大学, 1999年 - 2000年

    昆虫細胞-バキュロウイルス系による機能性タンパク質の高生産バイオリアクタープロセスを構築するための基礎を確立することを目的として,固定化していない細胞による組換えタンパク質の生産特性の解析および固定化による高密度培養と固定化細胞による組換えタンパク質生産について検討した. まず,組換えバキュロウイルスの感染時に昆虫細胞Sf9を新鮮培地に懸濁させ,種々の初期細胞密度および感染多重度(MOI)の下で振とう培養を行った.ウイルス感染時から組換えタンパク質の生産が完了する時刻までの生細胞密度の時間積分値に対して組換えタンパク質の生産量をプロットすることにより,組換えタンパク質生産に及ぼす細胞密度およびMOIの影響を解析した.組換えタンパク質生産は培地中の栄養分の有無によって著しく影響を受け,1以上の高いMOIでウイルスを細胞に感染させ,タンパク質生産の完了時に生細胞密度の時間積分値が8×10^6cells・d/cm^3に達するような初期細胞密度で培養を行うことにより,組換えタンパク質生産を至適化できることがわかった. 次に,多孔性の細胞保持粒子(biomass support particles,BSPs)を用いる昆虫細胞の固定化と組換えタンパク質生産について振とう培養において検討を行った.平均孔径が60μmのポリビニルホルマール樹脂多孔質体粒子(2×2×2mm)をBSPsとして用いると,Sf9は振とうフラスコ内で自然にBSPsに固定化され,定期的に培地交換しながら培養を継続すると,固定化細胞は5×10^7cells/cm^3-BSP以上の高密度に達することが示された.組換えウイルスを感染させた固定化細胞は,高密度状態においても,培地中の栄養分が枯渇しないように適切に培地交換しながら培養することにより,固定化していない細胞の最大比生産速度と同等の比速度で組換えタンパク質を生産可能であることが示唆された.

  • 動物細胞の低タンパク無血清培養と分離型バイオリアクターによる有用物質生産の効率化

    山地 秀樹

    日本学術振興会, 科学研究費助成事業 奨励研究(A), 奨励研究(A), 神戸大学, 1998年 - 1999年

    遺伝子組換えの宿主として現在広く利用されているChinese hamster ovary(CHO)細胞を対象にしたこれまでの検討から,ホスファチジン酸(PA)やリゾホスファチジン酸(LPA)などのリン脂質が無血清培養下におけるCHO細胞の増殖を促進することが示された.しかしながら,長鎖のアシル基を有するリン脂質は難溶性であり,培地に添加すると沈殿や凝集物の形成が認められることから,細胞にあまり有効に利用されていない可能性が示唆された.そこで本年度は,PAやLPAなどのリン脂質の細胞への効率的な供給方法の開発とこれらのリン脂質が組換えタンパク質生産に及ぼす影響について検討した. まず,リン脂質の新たな供給方法として,リン脂質をTween 80やPluronic F-68とともにマイクロエマルションを形成させる方法を検討した.その結果,リン脂質の溶解度が増大し,培地中での沈殿の形成も抑制できることがわかった.本法により調製したPAも無血清培養下におけるCHO細胞の増殖を促し,PA単独で添加した場合に比べて大きな細胞増殖促進効果が得られたことから,細胞がリン脂質をより有効に利用できるようになったものと考えられる. 次に,マイクロエマルション法により調製したPAやLPAを無血清培地に添加し,組換えCHO細胞の培養特性を調べた.これらのリン脂質は,組換え細胞の増殖を促進するだけではなく,細胞1個あたりの組換えタンパク質の生産量を増大させたことから,タンパク質生産を活性化する作用も有することがわかった.したがって,PAやLPAは,無血清培養において細胞増殖のみならず組換えタンパク質生産も促進する因子として活用できる可能性が示唆された.

産業財産権

  • ポリビニルホルマール樹脂多孔質体からなる微生物固定化用担体

    山地 秀樹

    特願2007-085105, 2007年03月28日, 大学長, 特許5069028, 2012年08月24日

    特許権