1994 年度実績報告書

下痢原因細菌の多種類の病原因子の同時迅速鑑別検出法の開発

研究課題

研究課題/領域番号	06557019
研究種目	試験研究(B)
研究機関	大阪大学
研究代表者	本田武司大阪大学, 微生物病研究所, 教授 (60029808)
研究分担者	飯田哲也大阪大学, 微生物病研究所・細菌感染分野, 助手 (90221746) 余明順大阪大学, 微生物病研究所・細菌感染分野, 助手 (70093482) 山本耕一郎大阪大学, 微生物病研究所・細菌感染分野, 助教授 (30158274)
キーワード	腸炎ビブリオ / 病原因子 / 耐熱性溶血毒 / 抗体 / ナイロン / 化学固定法
研究概要	1.新しく開発した化学的結合法(polyethlene imineとmaleic anhydride methylvinyl ether copolymerを用いた方法:PEI-MAMEC法)を用いて種々な高分子膜への抗体の結合量について検討したところ、Mylon膜が最も良かった。 2.腸炎ビブリオの病原因子である耐熱性溶血毒(TDH)およびその類似毒素(TRH)に対して特異的に反応するmonoclonal抗体を作成し、これをNylon小片に化学的に結合(固定)させた。 3.この両小片を一本のステックにセットし、未知検体(TDHかTRHのいずれかを含む)、抗TDH/TRH共通polyclonal抗体(ウサギ)、アルカリホスファターゼ標識抗ウサギ抗体、発色基質を順次反応させ、小片状の着色の有無でいずれかの毒素が検体中に存在するかを肉眼判定できる系を確立した。サンプル入手から結果判定までの全過程は一時間で完了した。従って、病原因子を迅速に同時識別検出するためのモデル系を開発することができた。 4.この系を動物実験系(腸炎ビブリオをchallengeしてウサギ結紮腸管ループ内貯留液を得、この液中のTDH/TRHの有無を判定)および旅行者下痢症の下痢便を用いた実用試験を実施し、予備的であるがpromisingな成績を得た。

研究成果
(12件)

すべてその他

すべて文献書誌 (12件)

[文献書誌] Xu,M.et al.: "Demonstration and characterization of simuvtaneous production of a TDH and a TRH by a clinical isolate of V.parahaemolyticus" Infection and Immunity. 62. 166-171 (1994)
[文献書誌] Tang,G.Q.et al.: "A mutant toxin of V.parahaemlyticus thermostable direct hemolysin which has lost hemolytic activity but retains ability to bind" Infection and Immunity. 62. 3299-3304 (1994)
[文献書誌] Xu,M.et al.: "Construction and characterization of an isogenic mutant of V.parahaemolyticus having a deletion inthe TRH gene." Journal of Bacteriology. 176. 4754-4760 (1994)
[文献書誌] Taniguchi,T.et al.: "Evidence that the N-terminal amino acid dequence of pilus colonization factor antigen III produced by human ETEC is similar to" Journal of Infectious Diseases. 170. 1049-1050 (1994)
[文献書誌] Ichinise,Y.et al.: "The effect on enterotoxicity of protease purified from Vibrio cholerae 01." FEMS Microbiol Letters. 115. 265-272 (1994)
[文献書誌] Miyagi,K.et al.: "Successful application of enzyme-1 beled oligonucleotide probe for rapid and accurate cholera-diagnosis in a clinical diagnosis" Microbiology and Immunology. 38. 301-304 (1994)
[文献書誌] Nagayama,K.et al.: "Correlation between Cell-associated mannose-sensitive hamagglutination by V.parahaemolyticus and adherence to a human colonic" FEMS Microbiology Letters. 120. 207-210 (1994)
[文献書誌] Shimaoka,M.et al.: "Development of enzyme-labeled oligonucleotide probe for mecA gene in MRSA." Journal of Clincal Microbiology. 32. 1866-1869 (1994)
[文献書誌] Iljima,Y.et al.: "Simultaneous outbreak due to Vibrio Cholerae and Shigella dysenteriae in Kenya." Lancet. 345. 69-70 (1995)
[文献書誌] Yoh,M.et al: "Evaluation of two assay kits for TDH as an indicator of TRH produced by V.parahaemolyticus." Microbiology and Immunology. 39. 157-159 (1995)
[文献書誌] Honda,T.et al.: "A novel method to chemically immobilize antibody on nylon and its application to the rapid and diferential detection of two toxins" Clinical and Diagnostic Laboratory Immunology. 2(in press). (1995)
[文献書誌] 本田武司: "病気と細菌毒素" 化学同人, 159 (1994)

1994 年度 実績報告書

下痢原因細菌の多種類の病原因子の同時迅速鑑別検出法の開発

研究代表者

本田 武司 大阪大学, 微生物病研究所, 教授 (60029808)

研究成果

[文献書誌] Xu,M.et al.: "Demonstration and characterization of simuvtaneous production of a TDH and a TRH by a clinical isolate of V.parahaemolyticus" Infection and Immunity. 62. 166-171 (1994)

[文献書誌] Tang,G.Q.et al.: "A mutant toxin of V.parahaemlyticus thermostable direct hemolysin which has lost hemolytic activity but retains ability to bind" Infection and Immunity. 62. 3299-3304 (1994)

[文献書誌] Xu,M.et al.: "Construction and characterization of an isogenic mutant of V.parahaemolyticus having a deletion inthe TRH gene." Journal of Bacteriology. 176. 4754-4760 (1994)

[文献書誌] Taniguchi,T.et al.: "Evidence that the N-terminal amino acid dequence of pilus colonization factor antigen III produced by human ETEC is similar to" Journal of Infectious Diseases. 170. 1049-1050 (1994)

[文献書誌] Ichinise,Y.et al.: "The effect on enterotoxicity of protease purified from Vibrio cholerae 01." FEMS Microbiol Letters. 115. 265-272 (1994)

[文献書誌] Miyagi,K.et al.: "Successful application of enzyme-1 beled oligonucleotide probe for rapid and accurate cholera-diagnosis in a clinical diagnosis" Microbiology and Immunology. 38. 301-304 (1994)

[文献書誌] Nagayama,K.et al.: "Correlation between Cell-associated mannose-sensitive hamagglutination by V.parahaemolyticus and adherence to a human colonic" FEMS Microbiology Letters. 120. 207-210 (1994)

[文献書誌] Shimaoka,M.et al.: "Development of enzyme-labeled oligonucleotide probe for mecA gene in MRSA." Journal of Clincal Microbiology. 32. 1866-1869 (1994)

[文献書誌] Iljima,Y.et al.: "Simultaneous outbreak due to Vibrio Cholerae and Shigella dysenteriae in Kenya." Lancet. 345. 69-70 (1995)

[文献書誌] Yoh,M.et al: "Evaluation of two assay kits for TDH as an indicator of TRH produced by V.parahaemolyticus." Microbiology and Immunology. 39. 157-159 (1995)

[文献書誌] Honda,T.et al.: "A novel method to chemically immobilize antibody on nylon and its application to the rapid and diferential detection of two toxins" Clinical and Diagnostic Laboratory Immunology. 2(in press). (1995)

[文献書誌] 本田武司: "病気と細菌毒素" 化学同人, 159 (1994)

1994 年度実績報告書

本田武司大阪大学, 微生物病研究所, 教授 (60029808)