| Research Abstract |
So far, almost all the cytometry techniques, involving (microscope-based) cytofluorometry and flow cytometry, have been made for the use of quantitative analysis of cellular content of bioactive materials (DNA, RNA, protein, etc.). In the proposed project, we developed a cytofluorometry system for tissue culture cells in vivo. The system is composed of a phase contrast microscope (NIKON TMD) equipped with both an epi-illumination photometer system and an auto-staging device, and of a personal computer (NEC PC-9801 Vm2). The computer programs (made by us with BASIC language) for both the data (dytofluorometry and cell positions) analysis and controlling the total system have been very important for carrying out this study. The data transfer between is done through RS-232C. The fluorescence staining for tissue dulture cells in vivo have also been investigated extensively (Hoechst 33342 for nuclear DNA, Fura 2 for free Ca ions, carboxyfluorescein diacetate-propidium iodide for viable and dead cells, fluorescence beads for phagocytic activity, etc.), and applied to the cytofluorometric studies. In particular, cell cycle analysis based on DNA content quantitation for the tissue culture cells in vivo could be carried out by determining the individual cellular positions and S-phase times in the cell cycle. Other cytofluorometric analysis (as for the above mentioned stainings), together with image processing, became also possible as an efficient means. In summary, the present cytofluorometry system appears to offer a new means for the functional and morphological analyses of the tissue culture cells in vivo, and to be important in the cell biology studies.
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