PLP-AMeX Method, Fixation Using PLP Fixative and Embedding in Paraffin by the AMeX Method, is Useful not only for Histochemistry but also In Situ Hybridization
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概要
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The authors have previously reported that the combination of fixation with periodate-lysine-paraformaldehyde (PLP) fixative and embedding in paraffin by the AMeX method (the PLP-AMeX method) resulted in better preservation of immunoreactivities and endogenous enzyme activities. In the present study, we investigated whether PLP-AMeX-processed tissue would be useful for in situ hybridization (ISH). The pancreases were removed from rats treated with or without streptozotocin (STZ), and was fixed in PLP fixative. After fixation, the specimens were processed and embedded in paraffin by the AMeX method. Paraffin sections were made on which ISH and immunohistochemical staining were performed. ISH was performed according to the non-radioactive method using digoxigenin-labeled oligodeoxynucleotides probes and anti-digoxigenin antibody. For ISH, a 28S rRNA antisense probe was used for evaluation of the level of hybridizable RNA in the sections. 28S rRNA was clearly detected in the various cells of the pancreas. Using an insulin antisense probe, the presence of insulin mRNA was clearly identified in the pancreatic islet-cells. Furthermore, optimal results for ISH of insulin mRNA and immunohistochemical staining of insulin could be obtained in consecutive sections from a PLP-AMeX-processed block. In the rats treated with STZ, the histological changes of pancreatic islet-cells and the changes in staining intensity of immunohistochemical staining for insulin protein and ISH for mRNA could be observed in the PLP-AMeX-processed sections. These findings suggest that hybridizable RNAs are well preserved by the PLP-AMeX method, and that the method is useful for ISH as well as immunohistochemical and enzyme histochemical analysis.
- 日本毒性病理学会の論文
- 2004-09-30
著者
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鈴木 雅実
Safety Assessment Dep. Fuji Gotemba Res. Laboratories Chugai Pharmaceutical Co. Ltd.
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WATANABE Takeshi
Department of Molecular Immunology, Medical Institute of Bioregulation, Kyushu University
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鈴木 雅実
Safety Assessment Department Chugai Pharmaceutical Co. Ltd.
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Suzuki Masami
中外製薬富士御殿場研究所
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Fujii Etsuko
中外製薬富士御殿場研究所
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Fujii Etsuko
Safety Assessment Department Chugai Pharmaceutical Co. Ltd.
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Fujii Etsuko
Toxicology Laboratory Chugai Pharmaceutical Co. Ltd.
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Fujii Etsuko
Department Of Veterinary Pathology Faculty Of Agriculture Iwate University
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TAKAI Hirotake
Department of Safety Assessment, Fuji Gotemba Research Laboratory, Chugai Pharmaceutical Co., Ltd.
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KATSUYAMA Kiyoka
Chugai Research Institute for Medical Science, Inc.
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SUZUKI Masami
Department of Safety Assessment, Fuji Gotemba Research Laboratory, Chugai Pharmaceutical Co., Ltd.
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杉本 哲朗
Fuji Gotemba Research Laboratories Chugai Pharmaceutical Co. Ltd.
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Sugimoto Tetsuro
Toxicology Laboratory Chugai Pharmaceutical Co. Ltd.
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Sakurai Takayuki
Safety Assessment Department Chugai Pharmaceutical Co. Ltd.
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Ito Tsuneo
Chugai Research Institute For Medical Science Inc.
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鈴木 雅実
Fuji Gotemba Research Laboratories Chugai Pharmaceutical Co. Ltd.
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Takai Hirotake
Safety Assessment Department Chugai Pharmaceutical Co. Ltd.
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SUGIMOTO Tetsuro
Department of Toxicology, Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd.
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TSUKAMOTO Kozue
Department of Toxicology, Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd.
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KATSUYAMA Kiyoka
Department of Toxicology, Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd.
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YOROZU Keigo
Department of Toxicology, Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd.
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KIMURA Kazuya
Department of Toxicology, Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd.
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ITO Tsuneo
Department of Toxicology, Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd.
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Tsukamoto Kozue
Department Of Toxicology Fuji Gotemba Research Laboratories Chugai Pharmaceutical Co. Ltd.
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Sugimoto Tetsuro
Department Of Pathology Keio University School Of Medicine
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Sugimoto Tetsuro
Safety Assessment Laboratory Chugai Pharmaceutical Co. Ltd.
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Katsuyama Kiyoka
Chugai Research Institute For Medical Science Inc.
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Kimura Kazuya
Department Of Biology Faculty Of Science Okayama University
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Suzuki Masami
Safety Assessment Department Chugai Pharmaceutical Co. Ltd.
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Ito Tsuneo
Safety Assessment Laboratory Chugai Pharmaceutical Co. Ltd.
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FUJII Etsuko
PharmaLogicals Research Pte. Ltd.
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Yorozu Keigo
Department Of Toxicology Fuji Gotemba Research Laboratories Chugai Pharmaceutical Co. Ltd.
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Watanabe Takeshi
Department Of Applied Biological Chemistry Faculty Of Agriculture Niigata University
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Suzuki Masami
Department Of Metallurgy
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Suzuki Masami
Department Of Biochemistry Faculty Of Veterinary Medicine Hokkaido University
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Watanabe Takeshi
Fuji Gotemba Research Laboratories Chugai Pharmaceutical Co. Ltd.
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Watanabe Takeshi
Department Of Agricultural Chemistry Faculty Of Agriculture Tohoku University
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SUZUKI Masami
Department of Agricultural Chemistry, Gifu University
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Kimura Kazuya
Department of Advanced Materials Science, University of Tokyo
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