Learning objectives
- Acquire knowledge of the principal techniques used for biomolecular analysis, with a focus on nucleic acids and proteins.
- Know how to interpret analytical data;
- Learn about the main applications in molecular diagnostics and chemical biology.
At the end of the course, the student is expected to:
- know the main biomolecular recognition processes underlying standard bioanalytical techniques and biosensors, including nucleic acid interactions, affinity binding processes and ligand-substrate interactions;
- have knowledge of the main biomolecular detection methods, including amplification techniques and immunoassays.
- be familiar with the main mass spectrometry-based instrumental techniques and the connected workflow used to analyze biomolecular species in complex samples.
- be able to read and process analytical data, also from a method validation perspective:
- recognize the potential, the limits and the drawbacks of the different techniques;
- develop critical thinking and creativity to address bioanalytical challenges
- build adequate communication skills and possess a good command of the terminology of the field.
- prove their ability to independently expand on the notions learned during the course, peruse the relevant literature and engage in the discussion of recent trends in bioanalytical chemistry.
Prerequisites
- - -
Course unit content
Principles of DNA nanotechnology and applications in bioanalytical chemistry: programmable dynamic interactions and mechanisms, enzymatic and non-enzymatic amplification techniques.
Immunoassays: enzymatic and fluorescence assays, complex/combined assays.
Principles of gel electrophoresis: separation of nucleic acids and proteins.
Mass spectrometry for biomolecular analysis: general principles, ion sources, mass analyzers, qualitative and quantitative acquisition modalities, tandem mass spectrometry, fundamentals of proteomics, hyphenated liquid chromatography-mass spectrometry techniques.
Data interpretation: principles of chemometrics, analytical parameters and method validation.
Laboratory: computer simulations, discussion and group presentation of recent articles on advanced bioanalytical methods.
Full programme
Principles of DNA nanotechnology and applications in bioanalytical chemistry: nucleic acid interactions and complex structures (duplex, triplex, G-quadruplex, i-motif, stem-loop hairpins, junctions, branched kissing loops); aptamers and their applications; artificial nucleic acids (PNA, LNA); dynamic DNA nanotechnology (strand displacement, toehold exchange, DNA switches, DNA walkers, effective molarity); enzyme-based amplification techniques (PCR, RT-PCR, LAMP, SDA, RCA, RPA, NASBA); enzyme-free amplification techniques (HCR, CHA).
Immunoassays: competitive/non-competitive; homogeneus/heterogenous; direct/indirect and sandwich ELISA; immunofluorescence; lateral flow assays; proximity ligation assays; agglutination assays.
Principles of gel electrophoresis: DNA separation on agarose and PAGE gels; protein separation on native and denaturing gels.
Mass spectrometry for biomolecular analysis: general principles; ESI and APCI ion sources; analyzers (quadrupole, TOF, ion trap, Orbitrap); tandem mass spectrometry MS/MS; qualitative and quantitative acquisition modalities; hyphenated LC-MS techniques; basic concepts of bottom-up proteomics; data dependent acquisition DDA; peptide fragmentation; MALDI; DESI.
Data interpretation: fundaments of chemometrics, analytical parameters and method validation (precision, trueness, dynamic range, LOD/LOQ, linear regression); calibration methods; matrix effect; analytical parameters in serological tests (specificity and selectivity, false negatives and false positives, Bayesian statistics, dissociation constants and binding affinity).
Laboratory: in silico modeling of DNA-based devices; computer-based exercises on statistics and method validation; group work: survey of recent trends bioanalytical chemistry and flash oral presentations of advanced methods (in English).
Bibliography
Lecture notes.
Articles from the literature recommended by the lecturer.
With regard to the mass spectrometry course section: “Mass Spectrometry: principles and applications”, E. de Hoffmann, V. Stroobant; Wiley, 2007.
For further insights: “Bioanalytical Chemistry”, S. R. Mikkelsen, E. Cortòn, Wiley, 2nd edition, 2016.
Teaching methods
Frontal teaching
Laboratory: computer simulations; group work (flash presentations in English).
Assessment methods and criteria
Oral exam
Other information
