1990 Fiscal Year Final Research Report Summary
Development of an Automated Dual Type of Micro Stopped-Flow/Flow-Injection Apparatus Applicable for Detection of Minute Alteration of Inorganic Phosphate or Phosphoric Group in Protein Level
| Project/Area Number |
01870077
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Research Field |
Functional basic dentistry
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| Research Institution | Okayama University |
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Principal Investigator |
TANIGUCHI Shigehiko Okayama University Dental School, Professor, 歯学部, 教授 (50034161)
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| Co-Investigator(Kenkyū-buntansha) |
YASUDA Kazuo Naka Works, Hitachi, Ltd., Superintendent Engineer, 計測事業部, 主管技師長
TAKAHASHI Kojiro Okayama University Dental School, Assistant Professor, 歯学部, 助手 (00144775)
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| Project Period (FY) |
1989 – 1990
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| Keywords | Hard tissues / Phosphorylation / Dephosphorylation / Micro-autoanalyzer / Stopped-flow method / Flow-injection analysis |
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| Research Abstract |
By combination of two sets of stopped-flow/flow-injection apparatus (single SF-FIA apparatus), an automated dual SF-FIA apparatus was constructed for serial measurements of minute alteration of inorganic phosphate (Pi) or phosphoric groups in protein level related to calcification of hard tissues or phosphorylation/dephosphorylation cycle of growth control system in normal/cancer cells. The dual SF-FIA apparatus was further installed by an intelligent auto sampler in order to carry out some complex pre-treatments of sample. Normally, the quatitative change of phosphorylation/dephosphorylation in protein level by phosphatase or kinase, respectively, can be detected spectrophotometrically through a direct method to free Pi using Malachite Green/molybdate reagent or an indirect method measuring NADH stoichiometrically reflecting ATP/ADP change in consecutive enzyme assay system. Furthermore, the dual SF-FIA apparatus could be applied as a rapid (msec order) stopping tool required for ^<32>P-labeling method by using filtration with milipore membrane. With 29.4 ul for each of sample and assay solution per one cycle measuring, half-hundred of serial measurements by using the dual SF-FIA apparatus are possible for the total volume of 3.0 ml for each of sample and assay solution. The time interval among the serial measurements with the dual SF-FIA apparatus can be set up within an arbitrary range longer than 4 sec.
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