| Project/Area Number |
02807108
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Psychiatric science
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| Research Institution | Niigata University, School of Medicine |
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Principal Investigator |
TOGASHI Shunji Niigata University, School of Medicine Department of Psychiatry Assistant, 医学部・附属病院, 助手 (30172141)
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| Co-Investigator(Kenkyū-buntansha) |
小林 徹 新潟大学, 医学部附属病院, 医員
増沢 菜生 新潟大学, 医学部附属病院, 医員
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| Project Period (FY) |
1990 – 1991
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| Project Status |
Completed (Fiscal Year 1991)
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| Budget Amount *help |
¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1991: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1990: ¥1,000,000 (Direct Cost: ¥1,000,000)
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| Keywords | Sigma receptor / Solubilization / Anti-idiotypic antibody / Immunoaffinity / Chromatography / Haloperidol / イムノ・ブロッティング |
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| Research Abstract |
I generated anti-idiotypic antibodies that specifically recognized the sigma receptor in the rat brain. Firstly, I immunized BALB/C mice with a haloperidol-bovine serum albumin conjugate and obtained monoclonal antihaloperidol antibodies (Anti-HAL). The anti-HAL recognized the piperidinyl moiety of haloperidol molecule. Secondarily, I immunized BALB/C mice with anti-HAL which were purified by protein A affinity chromatography and then coupled to keyhole limpet hemocyanin. Anti-idiotypic antibodies (anti-Id -mAb) which inhibited [ ^3H] -haloperidol binding to the anti-HAL were obtained. Sigma and D_2-dopamine receptors were assayed with [ ^3H] -haloperidol and [ ^3H] -spiperone, respectively. The anti-Id-mAb could inhibit [ ^3H] -haloperidol binding to the rat brain sigma receptor but not [ ^3H] -spiperone binding to D_2-dopamine receptors . Furthermore, anti-Id-mAb could immunoprecipitate only the sigma receptor from a detergent-solubilized preparation. In conclusion, the anti-Id-mAb produced in this study specifically recognized the sigma receptor.
The sigma receptor of the rat brain was solubilized with the detergent (1% digitonin and O. 1% cholic acid) and subsequently purified by immunoaffinity chromatography on anti-Id-mAb - Sepharose 4B. A single cycle of affinity chromatography resulted in a 300-fold purification. SDS-polyacrylamide gel electrophoresis of the partially purified sigma receptor showed the presence of a major band of Mr=30, 000 upon silver staining.
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