| Project/Area Number |
61440054
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| Research Category |
Grant-in-Aid for General Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Research Field |
Hematology
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| Research Institution | Kyushu University |
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Principal Investigator |
NIHO Yoshiyuki Kyushu University, Fac. Med., Professor, 医学部, 教授 (60091287)
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| Co-Investigator(Kenkyū-buntansha) |
KUDO Jiro Kyushu University, Fac. Med., Research Associate, 医学部, 助手 (90148940)
OKAMURA Takashi Kyushu University, Fac. Med., Research Associate, 医学部, 助手 (30136436)
SHIBUYA Tsunefumi Kyushu University, Fac. Med., Research Associate, 医学部, 助手 (70133171)
OKAMURA Seiichi Kyushu University, Fac. Med., Assistant Professor, 医学部, 講師 (20136435)
HARADA Mine Kyushu University, Fac. Med., Assistant Professor, 医学部, 講師 (00019621)
大塚 輝久 九州大学, 医学部, 助手 (20185317)
森岡 英次 九州大学, 医学部, 医員
大原 紀彦 九州大学, 医学部, 医員
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| Project Period (FY) |
1986 – 1989
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥18,100,000 (Direct Cost: ¥18,100,000)
Fiscal Year 1989: ¥5,100,000 (Direct Cost: ¥5,100,000)
Fiscal Year 1988: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1987: ¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 1986: ¥7,000,000 (Direct Cost: ¥7,000,000)
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| Keywords | granulopoietin / granulopoiesis / monoclonal antibody / G-CSF / GM-CSF / mass culture / enzyme-linked immunosorbent assay / CSF receptor / G-CSF / GM-CSF / G-CSF受容体 / GM-CSF遺伝子発現 / モノクローナル抗体 / GーCSF受容体 / GMーCSF遺伝子発現 / GーCSF遺伝子発現 / 大量細胞培養 / CSF遺伝子発現 / GMーOSF / 低分子ポリエチレングリコール / ELISA法 / 陰イオン交換クロマトグラフィー / 逆相クロマトグラフィー |
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| Research Abstract |
We have purified granulopoietin (colony-stimulating factor; CSF) from the conditioned medium (CM) of a CSF-producing cell line using anion exchange column chromatography, gel filtration column chromatography and reverse phase high pressure liquid chromatography. The final preparation showed a single band on SDS-polyacrylamide electrophoresis and possessed a specific activity of 10^8U/mg protein.
We then succeeded in establishing murine hybridomas secreting IgG reacting to human granulocyte CSF (G-CSF). Using this monoclonal antibody and the polyclonal antibody against G-CSF raised in rabbits, a very sensitive enzyme-linked immunosorbent assay (ELISA) was established. Similarly, the ELISA for granulocyte-macrophage CSF (GM-CSF) was also established after monoclonal and polyclonal antibodies against GM-CSF were made in our laboratory. The production of G-CSF or GM-CSF from fresh human cells and cell lines were studied using these sensitive assays. Serum levels of CSF were also studied using these ELISA and high serum levels were detected among patients suffering from infections, neutropenia, and some cases of acute myelogenous leukemia. We also studied their roles in vivo, CSFs were administered to mice and monkeys resulting in elavation of neutrophilic counts. CSFs were proven to be active in promoting clonogenic leukemic blast cell growth in vitro.
A significant number of the receptor for G-CSF was demonstrated in mature neutrophils, human cell lines of either myeloid or myelomonocytoid origins, leukemia blast cells from patients with acute myelogenous leukemia by radioreceptor assay. The cloning of EDNA encoding the receptor for G-CSF is now under the way.
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