| Project/Area Number |
06671596
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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 |
Urology
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| Research Institution | Ehime University |
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Principal Investigator |
IWATA Hidenobu Ehime University school of Medicine Associate Professor, 医学部, 助教授 (40108379)
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| Co-Investigator(Kenkyū-buntansha) |
YOKOYAMA Masayosi Ehime University school of Medicine Assistant Professor, 医学部附属病院, 講師 (50116993)
HAMADA Hitosi Ehime University school of Medicine Instructor, 医学部附属病院, 助手 (30243794)
ISEDA Tokuhiro Ehime University school of Medicine Instructor, 医学部, 助手 (50260386)
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| Project Period (FY) |
1994 – 1995
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| Project Status |
Completed (Fiscal Year 1995)
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| Budget Amount *help |
¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 1995: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1994: ¥1,100,000 (Direct Cost: ¥1,100,000)
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| Keywords | urinary protein / renal cell carcinoma / renal tubule / monoclonal antibody / tumor marker |
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| Research Abstract |
Urinary proteins originated from renal tubules were studied in order to find a possible tumor marker for renal cell carcinoma (RCC).
Urinary proteins were obtained from healthy adult male urine by ultrafiltration. Urinary proteins of plasma origin were removed by circulating the urinary proteins through an affinity column which contained immobilized rabbit anti-human whole serum immunoglobutins. Balb/c mice were immunized against the plasma protein-free urinary proteins. The spleen cells were fused with mouse myeloma cells. Hybridomas were screened by indirect immunofluorecence of fresh frozen tissue sections using culture supernatants.
Five hybridomas producing monoclonal antibodies (mAbs) which react with renal tubular segments, and two other hybridomas producing mAbs which react with basement membrane and connective tissue, and with urothelium were established. One mAb, named UP720, reacted strongly with distal renal tubules. Using this mAb, 39 RCCs were stained by immunoperoxidase method. Twenty-six of the 39 cancer tissues were stained positive. Therefore, the antigen recognized by UP720 (Ag720) can be a candidate for the tumor marker.
The estimated molecular weight of Ag720 was about 2OOkD by means of SDS-PAGE and immunoblottng.It was not stained by Coomassie blue, therefore, thought to be a sugar rich glycoprotein like a mucin. Immunoblotting of serum protein obtained from normal individuals and RCC patients failed to demonstrate the 2OOkD band. But serum Ag720 was detectable by sandwich ELISA.The purification of the antigen by an affinity column coupled with UP720 is fruitless until now.
The antigen recognized by an mAb which reacted with basement membrane and connective tissue was found to be alpha-1 microglobulin.
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