Reseach Of Confocal Microscopy For Space Life Science Experiments

Laser Scanning Confocal Microscopy (LSCM) has become an important tool formodern microscopy. Confocal ray path and fast scanning structure are used toacquire high resolution and3-D images in LSCM. Confocal microscopy in lifescience are widely used for quantitative analysis and location of thefluorescent-labeled molecular probes or structures in tissues and cells, real-timequantitative or semi-quantitative test of ion concentration, intercellularcommunication research, and determination of cell membrane fluidity. To meet theneeds of vivo confocal observation, this thesis carries out a study of rapidfluorescence confocal microscopy based on the low light detection and rapidscanning technology to acquire confocal fluorescence images of chlorophyll andGreen Fluorescence Protein (GFP).The aim of this thesis is to make the microscopyinstruments in space life science have high resolution imaging and sectioningcapability. It also can broaden the field of space life science experiments and providetechnical support for development of the space life science.The main content of this thesis is listed as following:1) The impact of different pinhole spacing and diameter on the system axialresolution is analyzed and the relationship between the pinhole diameter and axialresolution is calculated. The process of spatial sampling and the influence due tounder-sampling, the maximum pixel size without loss of the optical resolution andthe solution under the mismatch pixel size are analyzed. The two banding sourcesare located according to the scanning characteristics and the frame transfercharacteristics, then gives solution to reduce the impact of the banding.2) On target analysis and scheme comparison, EMCCD and pinhole disk arechosen for the rapid fluorescence confocal microscopy. The key components andtechnologies of system are analyzed and discussed, including the optics of imaging,exciting and fluorescence spectrum, pinhole disk structure and electronics.3) The rapid fluorescence confocal microscopy platform is setup, and theoptical system is calibrated. Identification plate is designed, and the calibration oflateral resolution is implemented. The relationship between pinhole diameter andaxial resolution is consistent with theoretical analysis according to the sectioning offluorophore target. Confocal images of plant leaves, Arabidopsis with GFP andquantum are captured.