Development of Wide-area Fluorescent Image Observing System for Analysis of the Relationship between Function and High-dimension Structure in Biological System.

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Recent fluorescent microscopy is working as an extremely powerful tool in biological science. Combining the newly developed fluorescent probes with an improved fluorescent microscope, we are now able to observe not only morphological structure, but also biophysical situation like intracellular distribution of Ca^<2+> and pH,etc. Furthermore, in plant cells, auto fluorescence of chlorophyll has been used to understand the various photosynthetic phenomena.In the present study, we planed to develop a new-type fluorescence observing system. This is an apparatus that equips a computer-controlled wide area irradiation system and high-sensitive video camera. Using this system, we will be able to observc a fluorescent imagc of wide area like a stereoscopic microscope.If we use newly developed fluorescent probes (such as BCECF,Fura 2, etcf.) with a wide-area fluorescence viewing systcm, we will be able to detect an interaction between cells in a tissue besides intracellular reactions in the individual cell.We basically planned the system and constructed the irradiation system and the fluorescent observing system in 1993. In 1994 and 1995, we improved the following points.(1) To detect the auto fluorescence of chlorophyll under saturated irradiation, the irradiation system with Xenon lump was improved. Furthermore, a halogen irradiation system was added to illuminate the sample.(2) Computer-controlled exchange system of filters was added to the irradiation system.(3) Video camera controlling system was improved to expand the detection range.(4) Computer program was developed for the fluorescent image analysis.(5) The irradiation system was improved to be applicable for the ordinal fluorescence microscope.(6) As the preliminary experiments, a fluorescent induction fo chlorophyll and non-photo chemical quenching was observed using this system.This apparatus is still under construction. We need further improvements of both hard and soft wares.