|
Lecturer(s)
|
-
Šebela Marek, prof. Mgr. Dr.
|
|
Course content
|
Introduction. Principles of working with biological material. Homogenization of tissues and extraction of cellular content. Extraction and precipitation methods. Centrifugation methods. Preparative and analytical centrifugation, centrifugation in density gradient. Dialysis. Ultrafiltration. Lyophilization. Chromatographic methods. Adsorption, ion-exchange, gel and affinity chromatography. HPLC and perfusion chromatography. Chromatofocusing. Electromigration methods. Gel electrophoresis. Isoelectric focusing and 2D electrophoresis. Blotting. Capillary electrophoresis. Methods of studying proteins. Assay of protein concentration, amino acid analysis, protein sequencing, peptide mapping, study of post-translational modifications, chemical modifications of proteins, stability studies, chemical synthesis of peptides. Mass spectrometry of proteins, peptide mass fingerprinting, de novo sequencing, characterization of posttranslational modifications, proteins as biomarkers in medicine. Analysis of nucleic acids. DNA mapping by restriction endonucleases, DNA chips, DNA sequencing, polymerase chain reaction (PCR), chemical synthesis of oligonucleotides. Immunochemical methods. Polyclonal and monoclonal antibodies. Immunochemical Immunodiffusion and immunoelectrophoresis. ELISA, immunoblotting. Flow cytometry. Spectrocopic methods. Absorption spectroscopy. Fluorescence spectroscopy. Infrared and Raman spectroscopy. Mösbauer spectroscopy. Chiroptic methods. Resonance spectroscopic methods - NMR a EPR. Protein crystallization. X-ray structural analysis. Electrochemical methods in biochemistry. Polarography and voltammetry. Biosensors. Microcalorimetry. Methods for determination of size and shape of biomacromolecules. Interaction of biomacromolecules with ligands. Study of fast reactions. Buffers, measurement of pH and conductivity. Enzyme analysis. Coupled reactions. Chromogenic and fluorogenic substrates. Radioactivity and its measurements. Autoradiography. Imaging.
|
|
Learning activities and teaching methods
|
Lecture
- Preparation for the Exam
- 75 hours per semester
- Attendace
- 39 hours per semester
|
|
Learning outcomes
|
Students become familiar with separation methods and instrumental analysis methods used in contemporary biochemistry.
Students will be able to define the main terms and describe the main experimental methods used in the biochemical laboratory.
|
|
Prerequisites
|
To understand the material, advanced orientation in the topics taught in the KBC/BCH Fundamentals of Biochemistry course is necessary.
KBC/BCH
|
|
Assessment methods and criteria
|
Written exam
Successfully passing the written exam - obtaining at least 50% of points. Evaluation: 20 - 18 p. A (1) 18 - 16.p B (1-2) 16 - 14 p. C (2) 14 - 12 p. D (2-3) 12 - 10 p. E (3)
|
|
Recommended literature
|
-
Anzenbacher P., Kovář J. (1986). Metody chemického výzkumu pro biochemiky. Praha.
-
Bhaskar, A. (2014). Biochemical Methods: A Practical Approach. Oxford.
-
Gault, V.A.; McClenaghan, N.H. (2009). Understanding Bioanalytical Chemistry: Principles and Applications. Chichester.
-
Hofmann, A., Clokie, S. (Eds.). (2018). Wilson and Walker´s Principles and Techniques of Biochemistry and Molecular Biology. Cambridge.
-
Hussain, K.; Lone, S.M.; Masoodi, K.Z.; Balkhi, S.M. (2024). Techniques for Biochemical Analysis. London.
-
Lottspeich, F.; Engels, J.W. (Eds.). (2018). Bioanalytics: Analytical Methods and Concepts in Biochemistry and Molecular Biology. Wiley-VCH.
-
Mikkelsen, S.R.; Cortón, E. (2016). Bioanalytical Chemistry. John Wiley & Sons, Hoboken, NJ, USA.
-
Mishra, N. Introduction to Proteomics: Principles and Applications. John Wiley & Sons, Hoboken, NJ, USA. 2010.
-
Pingoud, A.; Urbanke, C.; Hoggett J.; Jeltsch, A. (2002). Biochemical Methods : A Concise Guide for Students and Researchers.. Wiley-VCH.
|