| Project/Area Number |
04670890
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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 |
Orthopaedic surgery
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| Research Institution | Osaka University |
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Principal Investigator |
MASUHARA Kensaku Osaka University Medical School, Assistnat Professor, 医学部, 助手 (90238915)
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| Co-Investigator(Kenkyū-buntansha) |
ONO Keiro Osaka University Medical School, Professor, 医学部, 教授 (70028330)
YOSHIKAWA Hideki Osaka University Medical School, Assistnat Professor, 医学部, 講師 (60191558)
HASHIMOTO Jun Osaka University Medical School, Assistnat Professor, 医学部, 助手 (40237938)
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| Project Period (FY) |
1992 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1994: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1993: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1992: ¥1,100,000 (Direct Cost: ¥1,100,000)
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| Keywords | Monoclonal antibody / BMP-4 / Western blot analysis / Northern blot analysis / Immunohistochemistry |
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| Research Abstract |
A monoclonal antibody which reacts with murine and human bone morphogenetic protein-4 (BMP-4) has been developed using recombinant BMP-4 as an immunogen. The antibody that bound most tightly to recombinant murine (rm) BMP-4 was selected, subcloned and characterized. The specificity of the antibody was confirmed using Western blot analysis and enzyme-linked immunosorbent assay (ELISA). The antibody reacts with murine and human BMP-4 in both the reduced and non-reduced condition, however this antibody shows cross-reactivity with neither human BMP-2 nor TGI-beta1. Thus, the produced antibody could recognized the disulfide-linked dimeric structure of bioactive BMP-4, regardless of the species. Immunocytochemical study using this antibody successfully shows the cytosolic localization of BMP-4 in osteoinductive cells, i.e., BFO and Saos-2 in which the level of mRNA for BMP-4 was proved to be constitutively high by northern blot analysis. In addition, the antibody could demonstrate the presence of BMP-4 in developmental bone formation in the alveolar bone of rat embryo by immunohistochemistry. The antibody could be used for a more sensitive approach for quantitative analysis of BMP-4.
Sandwitch ELISA for quantification of BMP-4 in sera from patients with fractures and normal controls suggessted that BMP-4 in human serum could be below 1ng/ml.
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