2001 Fiscal Year Final Research Report Summary
Analysis of poly-N-acetyllactosamilie-bearing glycoproteins involved in neural cell differentiation
| Project/Area Number |
12672133
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biological pharmacy
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| Research Institution | Kyoto Sangyou University |
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Principal Investigator |
KUROSAKA Akira Kyoto Sangyou University, Faculty of Engineering, Professor, 工学部, 教授 (90186536)
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| Project Period (FY) |
2000 – 2001
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| Keywords | poly-N-acetvllactosamine / neurons / monoclonal antibodies / glycosyltransferases / in situ hybridization |
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| Research Abstract |
1. Preparation of monoclonal antibodies recognizing glycoproteins containing poly-N-acetyllactosamine carbohydrate chains.
(1) There are several glycoproteins containing poly-N-acetyllactosamine carbohydrate chains on the PC12 cell surface. In order to investigate expression of each glycoprotein, preparation of monoclonal antibodies was performed.
(2) Each glycoprotein was purified from a large-scale culture of PC12 cells by means of biochemical methods, such as gel filtration.
(3) Balb/c mice were immunized by injecting isolated glycoproteins intraperitoneally together with Freund's complete adjuvant.
(4) Although the antigenicity of the glycoproteins used for the immunization was low, significant increase in the blood liter was observed after repeated immunization of the mice.
(5) Hybridomas were prepared by fusing spleen cells from the immunized mice with myeloma cells. The screening of the hybridomas producing monoclonal antibodies specific for each glycoproteins is under progress.
2. Analysis of initial reaction of poly-N-acetyllactosamine biosynthesis.
(1) Expression of N-acetylgalactosaminyltransferases in the rat brain was analyzed by in situ hybridization, which catalyze the initial reaction of the biosynthesis of poly-N-acetyllatosamine carbohydrate chains.
(2) N-Acetylgalactosaminyltransferases were expressed specifically in the rat brain.
(3) They also exhibited characteristic expression in the restricted area of the brain, such as the intermediate layer of cerebral cortex, and hippocampus.
(4) With the production of specific monoclonal antibodies, it would be possible to study biosynthetic analysis of each glycoprotein by comparing the expression pattern in the brain between the N-acetygalactosaminyltransferases and the glycoproteins with poly-N-acetyllactosamine chains.
3. The results obtained were presented in the International Symposium on Glycoconjugates, and the annual meeting of Japanese Biochemical Society.
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