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Methods in Biochemistry

Teachers:
Paolo Iadarola, Gianni Francesco Guidetti
Year:
2017/2018
ECTS:
9
SSD:
BIO/10

Course content

Aim of the course is to provide students with the basic knowledge of techniques typically applied in a biochemical laboratory.


a) Methods for extraction and purification of proteins from animal/vegetal tissues and cultured cells. Procedures typically used in industry/research laboratories for primary purification of proteins. Theoretical principles and practical protocols of the most common chromatographic approaches: adsorption; partition; ion-exchange (IEC); gel-filtration (SEC); affinity; hydrophobic-interaction (HIC); perfusion; gas-chromatography (GC) and supercritical-fluid chromatography (SFC).Chromatofocusing; high performance liquid chromatography (HPLC); fast protein liquid chromatography (FPLC) and principles of green chromatography.
b) Electrophoretic techniques: analytical and preparative monodimensional (1-DE) electrophoresis of proteins in their native and denaturated state. Sodium-dodecyl-sulfate gel electrophoresis (SDS-PAGE). Isoelectrofocusing (IEF). Two-dimensional (2-DE) electrophoresis: application to proteomic studies. Capillary Zone electrophoresis (CZE) and Micellar Electrokinetic Chromatography (MEKC).
c) Spectroscopic techniques for the qualitative/quantitative characterization of proteins.
Adsorption spectroscopy: ultraviolet (U.V.); Visible; Infrared (IR); Nuclear Magnetic Resonance (NMR); Electron Spin Resonance (ESR). Emission spectroscopy: fluorescence and phosphorescence. Mass Spectrometry (MS).

Bibliography

Principles and Techniques of Practical Biochemistry by K. Wilson and J. Walker. 5th Edition. Cambridge University Press.


Credits: apnetwork.it