DESCRIPTION OF THE EDUCATIONAL ACTIVITY
Course title: Applied Biochemistry
Course number: 0730100134
Type of educational activity: Characteristic subject
Subject Group: BIO/10
Year of study: 2nd YEAR – Specialistic degree in Pharmacy
Total number of credits: 5
Global workload (n. of hours): 125
Number of hours allocated to: lectures, tutorials, laboratory, individual study: 40, 0, 0, 85
Teacher: LIBERATO MARZULLO
Objectives of the course:
Students will develop the following skills:
- to design operative schemes for the characterization and the study of molecules of
biological interest;
- to illustrate the theoretical principles underlying the most important methodologies used in
the biochemical studies.
Prerequisites:
Achievement of basic knowledges General biology, General Chemistry, Organic chemistry,
General and Applied Biochemistry.
Course contents:
Experimental systems in biochemical studies: in vivo and in vitro models.
Cell cultures. Microbial cultures. Cel count techniques.
Basic principles of FACS analysis and applications of flow cytofluorimetry.
UV-VIS spectroscopy: first elements, applications and instrumentation.
Spectrofluorimetry: first elements, applications and instrumentation.
Sedimentation and centrifugation: physical laws, preparative and analytical applications.
Preparation and fractionation of cell and tissue homogenates.
Assay methods for protein measuring.
Aminoacid composition and primary structure of proteins: analysis methods.
Protein fragmentation and peptide production: first elements of mass spectrometry in the
proteome analysis.
Electrophoresis: first elements and physical laws.
Electrophoresis in liquid and semisolid media: agarose gels and polyacrilamide gels.
SDS-PAGE and molecular mass determination of proteins.
Native PAGE.
Gradiente gels.
Isoelectrofocusing and protein pI.
Detection, evaluation and recovery of protein from gel.
Western and South-Western blotting.
Agarose and polyacrilamide gel electrophoresis of nucleic acids.
Pulsed field gel electrophoresis (PFGE)
DNA sequencing: Sanger and Maxam-Gilbert methods.
Chromatography: first elements and theoretical principles.
TLC [Link] low pressure chromatography. High performance liquid
chromatography (HPLC, FPLC).
Adsorption chromatography. Partition chromatography. Normal phase and reversed phase
(RPC) chromatography. Ion pair RPC. Hydrophobic interaction chromatography (HIC). Ion
exchange chromatography (IEX). Gel filtration chromatography. Affinity chromatography.
Working out of protein purification protocols.
Radiochemistry: nuclear reactions, radioisotopes, radioactive decay.
Matter-radiation interactions: safety elements in radioprotection.
Detection and measuring of radioactivity: autoradiography, gas ionization methods, scintillation
methods.
Practical aspects in radioactivity counting and data analysis.
Uses of radioisotopic tracers in biochemical, biomolecular and biomedical analyses.
Immunochemistry: monoclonal (mAb) and polyclonal (pAb) antibodies production.
In vivo and in vitro radiolabeling of proteins and nucleic acids.
Purification and fragmentation of immunoglobulins.
Immunoprecipitation.
Antibody labeling. Immunoassays: RIA, IRMA, ELISA.
Nucleic acids analysis:Northern and Southern analyses, restriction enzymes, modification
enzymes and their applications.
Polymerase chain reaction (PCR). Sensitivity and general applications of PCR.
Cloning vectors and molecular cloning.
Production of recombinant proteins.
Recommended text books:
Wilson, Keith & Walker, John -Principles and Techniques of Biochemistry and Molecular Biology
- 6th ed. Cambridge University Press, 2005
(Italian edition: WILSON Keith & WALKER, John - Biochimica e Biologia Molecolare: Principi e
Tecniche - Raffaello Cortina ed. (ISBN 88-6030-066-5))
Suggested readings: //
Teaching methods: lectures
Assessment methods: final oral examination
The course is taught in Italian
Additional information: further information can be requested by e-mail: marzullo@[Link]
