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Lecturer(s)
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Řehák Jan, Mgr.
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Luptovčiak Ivan, Mgr. Ph.D.
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Ovečka Miroslav, prof. Mgr. Ph.D.
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Hlaváčková Kateřina, Mgr. Ph.D.
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Course content
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1. Principles of light microscopy. Features of the light and functions of optical lenses. The construction of the optical elements of the light microscope and imaging principle. Distribution and basic types of light microscopes, the parameters utilized in the research of plant cells 2. Types of microscopic objects, methods of their preparation: native, permanent, whole objects and sections. Fixation, microscopic dyes, histochemistry and cytochemistry. 3. The construction of microscopic objectives, the theory of image formation and magnification. Types of objectives and possibilities of their application in different types of imaging of plant samples. 4. Special techniques of light microscopy I. Contrast in the light microscope, dark-field imaging, the principle of phase contrast and polarization. 5. Special techniques of light microscopy II. The principle of differential interference contrast and other methods of modulation contrast in light microscopy. 6. Principle of fluorescence microscopy. Construction of the fluorescence microscope, theory of the fluorescence filters, the use of fluorochromes. 7. Application of fluorescence microscopy in studies of plant cells. Autofluorescence, secondary fluorescence, immunofluorescence, fluorochromes for in vivo microscopy. 8. Methods of studying plant cells using fluorescent proteins. 9. Confocal laser scanning microscopy. The principle of optical sections, methods of scanning in the confocal microscope. Specialized methods of confocal microscopy and their use for advanced analysis of samples. 10. Advanced microscopic methods. Vital microscopy: microscopes to monitor fast cellular processes, videomicroscopy, live cell imaging. High resolution microscopy: super-resolution and recording of the fluorescence of single molecules. 11. Documentation and image analysis of microscopic images. Sensor device for detecting the transmitted light and fluorescence. Electronic signal adjustement. The graphic layout of the data and their use. 12. Fundamentals of electron microscopy. Principles of sample preparation, applications in biotechnology in comparison with light and fluorescence microscopy.
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Learning activities and teaching methods
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Lecture, Dialogic Lecture (Discussion, Dialog, Brainstorming), Demonstration
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Learning outcomes
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Comprehensive training of students on theoretical and practical solutions of the fundamental problems of plant studies at the microscopic level
Students will acquire basic theoretical knowledge about the range of microscopic techniques and practical experiences with their applications in the study of plant cells, tissues and organs.
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Prerequisites
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unspecified
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Assessment methods and criteria
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Written exam
Participation at the practical part and preparation of the protocols Successful completion of a written examination test form with a minimum of 75% of earned points
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Recommended literature
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Cox G. (2012). Optical imaging techniques in cell biology. CRC Press, Boca Raton .
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Goldman, R. D., Swedlow, J. R., & Spector, D. L. (2010). Live cell imaging : a laboratory manual . Cold Spring Harbor, N.Y: Cold Spring Harbor Laboratory.
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Hejtmánek M. (1994). Praktická cvičení z biologie. UP, Olomouc.
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Hejtmánek, M. (2001). Úvod do světelné mikroskopie. V Olomouci: Univerzita Palackého.
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Price RL, Jerome WG. (2010). Basic confocal microscopy. Springer, Heidelberg.
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Ruzin SE . (1999). Plant microtechnique and microscopy. Oxford University Press, Oxford.
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Spector, D. L., & Goldman, R. D. (2006). Basic methods in microscopy : protocols and concepts from cells : a laboratory manual. New York, N.Y: Cold Spring Harbor Laboratory Press.
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