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Lecturer(s)
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Berka Karel, prof. RNDr. Ph.D.
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Hof Martin, prof. Dr. Ph.D.
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Course content
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Main attention is paid to the fundamental findings in the field of quot, steady-steady quot and time differentiated fluorescent spectra, theory of their formation and their interpretation. Theoretical introduction is completed by the practical example of not only typical analytical applications but also of the latest usage of these methods in the study of the characteristics of the biological systems and chemical systems - micro-viscosity, relaxation times of the solvent layers, study of structural changes of proteins. Syllabus: 1. Influence of quenching on fluorescence quantum yield and lifetime. 1.1. Fluorescence quantum yield and lifetime. 1.2. Fluorescence quencher.1.2.1 Solute quenching 1.2.2 Example for application of solute quenching in protein studies 1.2.3 Solvent quenching 1.2.4 Self quenching 1.2.5 Trivial quenching 2 Influence of solvent relaxation on steady-state and time-resolved fluorescence spectra 2.1 Basic principles of solvent relaxation 2.2 Influence of solvent relaxation on steady-state spectra 2.2.1 Non-viscous solvents 2.2.2 Viscous and virtified solutions 2.3 Quantitative characterisation of solvent relaxation by time-resolved spectroscopy 2.3.1 Time-resolved emission spectra 2.3.2 Example for using solvent relaxation for probing micro-polarities 3 Information included in polarized fluorescence 3.1 Introduction 3.2 Definition of polarization and anisotropy 3.3 Steady-state fluorescence anisotropy 3.4 Time-resolved fluorescence polarization 4 Using fluorescence resonance energy transfer as a spectroscopic ruler 4.1 Donor-acceptor pairs at fixed distances 4.2 Donor-acceptor pairs at variable distances 4.3 Some applications of fluorescence resonance energy transfer 5 Information gained from excimer formation studies 6 Utilization of irreversible photobleaching 7 Single molecule detection by fluorescence
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Learning activities and teaching methods
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Dialogic Lecture (Discussion, Dialog, Brainstorming)
- Preparation for the Exam
- 0 hours per semester
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Learning outcomes
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The lectures are aimed on the introduction into fluorescent spectroscopy and related methods.
ability to evaluate particular methods and procedures, explain results related to fluoroscent spectroscopy
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Prerequisites
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unspecified
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Assessment methods and criteria
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Oral exam
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Recommended literature
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Ganglitz G. (ed.). (1998). Handbook of Spectroscopy. Weinheim.
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Lakowicz, J. (1999). Principle of fluorescence spectroscopy. Kluwer.
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Prosser, V. a kol. (1989). Experimentální metody biofyziky. Academia Praha.
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Slavík, J.. (1994). Fluorescent Probes in Cellular and Molecular Biology. CRC Press.
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