2000 Fiscal Year Final Research Report Summary
Highly-Selective Determination of Chemical Sepecies in Cells with 3D Fluorescence Microscopy
| Project/Area Number |
10555294
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
工業分析化学
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
HARATA Akira KYUSHU UNIVERSITY, Faculty of Matrl. Sci., Ass. Prof., 大学院・総合理工学研究院, 助教授 (90222231)
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| Project Period (FY) |
1998 – 2000
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| Keywords | Laser-Induced Fluorescence / Fluorescence Microscope / Spectroscopic Image / Three Dimensional Analysis / Highly-Selective Determination / Biological Substances / Ion Exchange Resin / Particle Counting |
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| Research Abstract |
A fluorescence method is the one of the most sensitive detection tools for chemical species but determination of amounts of functional substances in a biological cell for diagnostics always faces many practical problems : Large backgrounds caused by nonspecifically adsorbed-fluorescence probe and organic substances makes high sensitive detection impossible ; Light scattering from the cell walls disturbs reproducibility ; and so on. This study aims to solve these problems and get practical guidelines for a fluorescence method of cell-based diagnostics.
1) A three-dimensional fluorescence micro-spectrometer is composed. A laser source or monochromated halogen-lamp light source, a microscope and an imaging spectrometer are combined to observe spectral images of microscopic substances.
2) Discrimination of individual microparticles is demonstrated with the fluorescence micro-spectrometer. Different types of fluorescence dyes are adsorbed on different microparticles, respectively. They are made of polystyrene or ion-exchange resin, and their diameters are from 3 to 11 μm. After mixing, they are used as test samples. It was found that each microparticle was distinguishable even if their fluorescence emission spectra have the same shapes with a difference in their peak wavelengths of only 5-10 nm. Particle discrimination under a high background condition was demonstrated as well.
3) Detection limit for adsorbed dyes on one microparticle was determined to be as low as 10^<-18> mol/particle.
4) Based on the experimental results, a guideline for fluorescence determination of chemical species in micro-substances was proposed where fluorescence intensity, spectra and background levels are evaluated for the diagnostics.
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