91309 - Medical Applied Biochemistry

Learning outcomes

At the end of the course, the student will have learned:
- the main biochemical techniques and methodologies of biochemical analysis
- the rationale behind the applications of these technologies to medicine and the clinic;
- how to apply biochemical methods to the clinical analyses, to understand and correctly interpret the results also from a diagnostic/prognostic point of view;
- communicate to patients the correct reading of reports of simple biochemical-clinical analyses.

Course contents

UV-VIS Absorption Spectroscopy. Physical principles of absorption of electromagnetic radiation. Lambert-Beer's law. Instrumentation and applications. Chromophores in biomolecules. Concentration determination of proteins and nucleic acids. Fluorescence spectroscopy. Physical principles and instrumentation. Extrinsic and intrinsic fluorophores. Fluorescence polarization. FRET. Chemiluminescence, bioluminescence. Luminometry and applications. Principles of cytofluorimetry and its applications.

Chromatographic techniques. General principles. Ion exchange chromatography. Molecular exclusion chromatography or gel filtration. Affinity chromatography. Partition chromatography. Adsorption chromatography. Applications of the techniques. Purification of proteins.

Centrifugal sedimentation techniques. Preparative and analytical applications. Hematocrit.

Electrophoresis. Principles of the technique. SDS-PAGE. Isolectrofocusing. gel-electrophoresis. Blue Native protein gel electrophoresis. Electrophoresis and analysis of nucleic acids. Applications of biomedical relevance.

Immunochemistry. General principles. Antibody structure and antigen interactions. Polyclonal and monoclonal antibody production. Precipitation of Ab-Ag complex (equivalence zone). Simple and double immunodiffusion techniques. Immunoelectrophoresis. ELISA, immunofluorescence, DELFIA, RIA. Immunodetection of separated proteins by electrophoresis (WB). Analytical and diagnostic applications.

Radiochemical techniques. General principles. RIA: Radio-Immuno-Assay. In vivo diagnostics. Scintigraphic techniques. PET.
Molecular Biology Techniques. Restriction enzymes and DNA analysis. Hybridization and transfer techniques on cellulose supports (blotting). Fingerprinting and Polymorphisms. Clinical applications: Diagnosis of hereditary diseases. DNA Microarrays. PCR, principles and applications.
Cell cultures, principles and applications.

 

Laboratory (2 CFUs)

Assay of protein content in biological samples: Bradford method. Set up of calibration curve and determination of protein content of an unknown sample.
Spectrophotometric assay of enzyme activity, construction of a Michaelis-Menten curve and determination of KM and Vmax values.
Determination of antioxidant power by FRAP assay. Preparation of calibration curve and determination of antioxidant power in different compounds.
Determination of cell viability.
Cell counts and staining of cell structures with hematoxylin/eosin.

Readings/Bibliography

• Metodologie biochimiche e biomolecolari. Mauro Maccarone Ed. Zanichelli
• Biochimica Clinica. Luigi Spandrio Edizioni Sorbona
• Biochimica Applicata, autori Stoppini-Bellotti Editore EdiSES, Napoli
• Metodologia biochimica - Le bioscienze e le biotecnologie in laboratorio A cura di Keith Wilson e John Walker Edizioni Raffaello Cortina Editore, Milano

Teaching methods

The course includes 4 frontal credits, in which a series of biochemical techniques and their medical application will be considered, and 2 laboratory credits in which problems such as: determination of the protein content of a solution, determination of enzymatic kinetic parameters, cell counting or staining of cellular samples will be addressed from a practical point of view. The presentations of the lectures will be deposited in the Virtual Learning Platform.

 

As concerns the teaching methods of this course unit, all students (including all the international incoming exchange students, i.e. ERASMUS) must attend Module 1, 2 online [https://www.unibo.it/it/servizi-e-opportunita/salute-e-assistenza/salute-e-sicurezza/sicurezza-e-salute-nei-luoghi-di-studio-e-tirocinio], while Module 3 on health and safety is to be attended in class. Information about Module 3 attendance schedule is available on the website of your degree programme ("studiare"--"formazione obbligatoria su sicurezza e salute").

Assessment methods

In order to take the exam, it is necessary the registration on the almaesami website, respecting the deadlines. Those who are not able to enroll by the due date, are required to communicate promptly (and in any case before the official closure of the registration lists) the problem to the teacher by e-mail.The learning assessment will be done through the final oral exam. The student will discuss a short elaborate concerning the application of one of the techniques covered during the course and general questions about the techniques considered during the course. The evaluation will be based on the answers given to each question and the ability demonstrated to have learned the fundamental notions to achieve the educational objectives. The definition of that knowledge and its critical mastery will contribute to the final grade. The student will also be required to submit a report on the laboratory, which will be evaluated. To take the exam, you must have attended the course: "Biochimica generale, molecolare e metabolica".

Teaching tools

 

Virtuale platform

Office hours

See the website of Christian Bergamini