| Project/Area Number |
60480280
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Hematology
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| Research Institution | Nagoya University School of Medicine |
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Principal Investigator |
SAITO Hidehiko Nagoya University School of Medicine, 医学部, 教授 (20153819)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAMATSU Junki Nagoya University School of Medicine, 医学部, 医員
KAMIYA Tadashi Nagoya University School of Medicine, 医学部, 助手 (50111844)
杉原 卓朗 名古屋大学, 医学部, 医員
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| Project Period (FY) |
1985 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥5,300,000 (Direct Cost: ¥5,300,000)
Fiscal Year 1987: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1986: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1985: ¥3,500,000 (Direct Cost: ¥3,500,000)
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| Keywords | Hypercoagulability / Activated factor IX / Antithrombin III / 活性【X】因子-アンチトロンビン【III】複合体 / 活性【IX】因子-アンチトロンビン【III】複合体 |
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| Research Abstract |
An enzyme-linked immunosorbent Assay (ELISA) using activated Thiol-Sepharose has been developed for the quantitation of activated human factor IX-antithrombin III (IXa-AT III) complex. IXa-AT III complex is bound to a mini-column of thiol-Sepharose to whixh monospecific rabbit antibody against AT III was coupled. IXa-AT III complex is then quantified by a second antibody, rabbit anti-human factor IX Fab', labeled with <beta>-d-galactosidase. IXa-AT III complex was detected, when a final concentration of 5.0 ng[1.0 ug/ml of purified IXa-AT III complex was added to buffer or plasma. The minimum detectable amount of the complex (5 ng/ml) corresponds to activation of as little as 0.05% of plasma factor IX. Experiments with purified F.IXa, AT III, normal plasma and AT III-depleted plasma demonstrated the specificity of the assay. We measured Ixa-At III complex generation in clotting normal blood in glass tubes, and found that IXa-AT III complex appears within 60 sec after venipuncture. No detectable amount of the complex was observed in native whole blood from patients complex in 10 normal individuals were less than 5.0 ng/ml, wheresa those in 3 patients with disseminated in travascular coagulation (DIC) were more than 20 ng/ml. The ELISA using activated thiol-Sepharose described in this paper seems to provide a useful tool to detect activation of coagulation system in vitro as well as in vivo.
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