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Lecturer(s)
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Vancová Marie, RNDr. Ph.D.
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Weyda František, doc. RNDr. CSc.
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Sehadová Hana, Mgr. Ph.D.
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Fessl Tomáš, Mgr. Ph.D.
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Bondar Alexey, Mgr. Ph.D.
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Krejčová Gabriela, Mgr.
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Course content
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Content of lectures: 1) Introduction - history of microscopy, optic apparatus, types of objectives, types of microscopes 2) Classical light microscopy + Keller illumination and techniques of contrast - Nomarski contrast (DIC), Hoffman contrast (HMC), phase contrast, dark field 3) Fluorescence microscopy - principle, overview of fluorophores, types of microscopes, application 4) Confocal microscopy - principle, types of microscopes, application - FRAP, FRET, multiphotone and TIRF microscopy 5) Superresolution techniques - PALM, STORM, STED, SIM, Airy scan 6) Luminescence microscopy - principle, application 7) Detection of the light signal - CCD and EM-CCD cameras, photomultipliers, clarification of the HDR and deconvolution 8) Insight into the electron microscopy (Ing. Jana Nebesářová, CSc.) 9)Techniques of macro photography (Doc. RNDr. František Weyda, CSc.) 10) Image analysis_1 - software Adobe Photoshop 11) Image analysis_2 - software FIJI (ImageJ) 12) Image analysis_3 - software IMARIS a XuvStitch Content of practicals: 1) Practical demonstration of the microscope adjustment (Keller illumination, Nomarski contrast) - (Microscopy lecture room) 2) Demonstration of the fluorescence microscope - (Bioscience Imaging and Histology Unit) 3) Demonstration of the confocal microscope - (Bioscience Imaging and Histology Unit) 4) Excursion to the Olympus company (Prague) or demonstration of the microscopes in the University) 5) Demonstration of the luminescence microscope - (Bioscience Imaging and Histology Unit) 6) Excursion to the Laboratory of electron microscopy 7) Demonstration of macro photography taking 8) Image analysis in the Adobe Photoshop software_part 1 - (Computer lecture room) 9) Image analysis in the Adobe Photoshop software_part 2 - (Computer lecture room) 10) Image analysis in the Adobe Photoshop software_part 3 - (Computer lecture room) 11) Image analysis in the FIJI (ImageJ) software - (Computer lecture room) 12) Demonstration of the IMARIS software - (Bioscience Imaging and Histology Unit)
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Learning activities and teaching methods
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Monologic (reading, lecture, briefing), Dialogic (discussion, interview, brainstorming), Monitoring, Demonstration, Excursion, Work with multi-media resources (texts, internet, IT technologies), Practical training
- Class attendance
- 50 hours per semester
- Preparation for classes
- 30 hours per semester
- Field trip
- 5 hours per semester
- Preparation for credit
- 20 hours per semester
- Preparation for exam
- 40 hours per semester
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Learning outcomes
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Students will learn the principles of basic microscopy techniques and gain knowledge about the image analyses of biological material usable for scientific publication.
Students will learn who manipulate the bright-field, fluorescence, and confocal microscope, bioluminescence microscope and the principles of superresolution and electron microscopy. They become also acquainted with the less common microscopes by several excursion and thus they will obtain a comprehensive idea about the recent microscopy. They will be train how to adjust the light pathway in different types of microscopes to be able independently manipulate any type of bright-flied and fluorescence microscopes. Students should be able to choose the best method for visualization of variable biological objects. They will learn the basic common software for digital image analysis such are Adobe Photoshop and ImageJ.
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Prerequisites
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The course introduces the issue, does not assume specific input knowledge.
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Assessment methods and criteria
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Oral examination, Student performance assessment, Systematic student observation, Test
Requirements for credit: 1) presence on the practical trainings (maximum one absence) 2) independent adjustment of the bright-field and fluorescence pathway in a microscope 3) successful passing of final test Requirements for oral exam: 1) obtaining of the credit
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Recommended literature
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http://olympus.magnet.fsu.edu/primer/techniques/confocal/applications/fretintro.html.
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http://olympus.magnet.fsu.edu/primer/techniques/confocal/index.html.
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http://olympus.magnet.fsu.edu/primer/techniques/fluorescence/fluorhome.html.
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https://www.microscopyu.com/techniques/confocal.
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https://www.microscopyu.com/techniques/fluorescence.
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Nathan S. Claxton, Thomas J. Fellers, and Michael W. Davidson Laser Scanning Confocal Microscopy.
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