Lecture
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18 Nov 2021 (1h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Presentazione corso.
22 Nov 2021 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
experimental uncertainties, probability distributions, error bars, hints on statistical tests; statistical estimators, error propagation (beginning).
01 Dic 2021 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Error propagation (continued); covariance matrix and correlation. Noise in measurements, examples in spectroscopy. Fundamentals of lenses. Microscopy: objectives, resolution, numerical aperture.
06 Dic 2021 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Electromagnetic radiation, refraction and diffraction of light, limit of diffraction for resolution; optical path in transmission microscopes, conjugate planes, field diaphragm and aperture. Optical paths in reflection / epifluorescence microscopes. Transmission microscopy: Köhler illumination, contrast techniques (introduction, polarization contrast, dark field); optical path and phase differences.
06 Dic 2021 (2h 00m)
Francesco Cardarelli - Didattica integrativa - Mista
Lezione 1
13 Dic 2021 (1h 00m)
Francesco Cardarelli - Didattica integrativa - Mista
Lezione 2
15 Dic 2021 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Contrast techniques (phase contrast, DIC). Alignment of a transmission microscope. Contrast techniques in reflection microscopy. Epifluorescence microscopy. Outline of fluorophores (and immunofluorescence) and dichroics / optical filters. Confocal microscopy (introduction)
15 Dic 2021 (1h 00m)
Francesco Cardarelli - Didattica integrativa - Mista
Lezione 3
20 Dic 2021 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Confocal microscopy: implementations, Point Spread Function, convolution; magnification and resolution; dependence on signal/noise ratio, variable with laser intensity and detector gain. Dependence of signal/noise ratio and of resolution on the confocal pinhole aperture.
20 Dic 2021 (1h 00m)
Francesco Cardarelli - Didattica integrativa - Mista
Lezione 4
10 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Dependence of signal/noise ratio and of resolution on the confocal pinhole aperture (recall and completion). Aberrations (spherical, chromatic). 2 photon microscopy; TIRF microscopy. Fluorescence microscopy: labelling and types of fluorophores (fluorescent proteins, Qdots, organic fluorophores); chemical tags (ACP and its derivatives): methods of insertion into proteins (hints on transfection, vectors, plasmids, promoters, controlled expression). Advanced methods in fluorescence microscopy: colocalization.
10 Gen 2022 (1h 00m)
Francesco Cardarelli - Didattica integrativa - Mista
Lezione 5
12 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Advanced methods in fluorescence microscopy: quantifications of colocalization; FRET and measurement methods (sensitized emission, acceptor photobleaching). FLIM (hints). FRET-FLIM with examples. Laws of diffusion.
17 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Fluorescence microscopy techniques: laws of diffusion (cont.), sum over microtrajectories; FRAP and other fluorescence microscopy techniques based on photobleaching: analytical treatment (diffusive model, compartment model), experimental details (corrections and normalizations). FLIP; iFRAP, photoactivation (PA), FLAP, photoconversion (PC). FCS (introduction).
19 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
FCS. Spectroscopy of single molecules (outline and techniques); Single molecules tracking: analysis; types of trajectories, MSD; anomalous diffusion, transient arrest of diffusion. Hints on experiments on membrane receptors and neurotrophins: 2D-distributions of couple of parameters, counting fluorescent molecules in tracked spots. Deconvolution.
24 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Light-matter interaction: generalities. Quantum mechanics review; time-dependent perturbation theory, Fermi's golden rule. Review of Fourier series and transform, Taylor series; Dirac delta.
26 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Interactions with radiation. Einstein coefficients; operating principles of lasers. Jablonski diagrams and absorption. Electronic and vibrational spectroscopy: Born Oppenheimer approximation, IR absorption.
31 Gen 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Vibrational modes and selection rules. Vibronic transitions. Franck Condon factors. Properties, setups and measurement methods for optical absorption (UV-visible-NIR); spectrometers and grating. Lambert-Beer law.
02 Feb 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Fluorescence properties, and measurements methods and setups. Solvent relaxation in fluorescence. Spectrofluorimeter interface. Non linear light-matter interactions. Differences amongst 2-photon absorption and second harmonic generation. Second order perturbation theory, 2 photon absorption, Raman.
07 Feb 2022 (1h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Raman: Stokes/antiStokes intensity ratio, thermally activated fluorescence. Vibrational spectroscopy (also setups): IR absorption (outline) and Raman.
09 Feb 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Vibrational and Raman modes in fluorescent proteins (continued). SERS and surface resonance plasmons in metal nanoparticles. Organic fluorophores: orbitals (pi, sigma, non-bond).
16 Feb 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - A distanza
Organic fluorophores: molecular transitions; electronic delocalization. Molecular states of formaldehyde. Molecular transitions in formaldehyde (outline). Examples of organic fluorophores; notes on immunofluorescence and the use of organic fluorophores in fixed or living cells and in vivo. AM loading. Fluorescent quantum dots: structure, spectra, functionalizations.
23 Feb 2022 (2h 00m)
Riccardo Nifosì - Didattica integrativa - Mista
Molecular forces. Structure of carbohydrates, lipids and nucleic acids.
28 Feb 2022 (2h 00m)
Riccardo Nifosì - Didattica integrativa - Mista
Structure and dynamics of proteins: amino acids and their properties; post-translational modifications; peptide bond; secondary, tertiary and quaternary structure; Ramachandran map; stability, denaturation and folding.
02 Mar 2022 (2h 00m)
Riccardo Nifosì - Didattica integrativa - Mista
Methods for the determination of protein structure: X-ray crystallography and NMR. Protein data bank. Computational modelling of biomolecules: multi scale models; molecular mechanics and molecular dynamics.
09 Mar 2022 (2h 00m)
Riccardo Nifosì - Didattica integrativa - Mista
Computational methods for protein structure prediction and protein design. Fluorescent proteins: structure and chromophore formation; optical properties; applications.
23 Mar 2022 (2h 00m)
Stefano Luin - Corso (attività didattica) - Mista
Transitions in a 2-level system; life times, measurement methods (photobleaching rate, frequency and time domain measurements); multiexponential decay; temporally resolved microscopy / FLIM; instrumental considerations (streak camera, gated acquisition, TCSPC, IRF, quadrant mode microchannel plates). Phasor method in FLIM. Superresolved microscopy: Abbe limit; 4Pi microscopy; STORM and FPALM (general information, localization precision, tools); RESOLFT and STED (Concept and generality, saturation, stimulated emission, instrumentation, engineering of the "extinguishing" PSF), photoconversion vs "emptying" of the excited state, typical times; comparisons between RESOLFT and STORM / FPALM.
09 Maggio 2022 (2h 00m)
Gian Michele Ratto - Corso (attività didattica) - In presenza
Imaging a due fotoni in vivo: imaging anatomico funzionale in vivo.
11 Maggio 2022 (2h 00m)
Gian Michele Ratto - Corso (attività didattica) - In presenza
Imaging a due fotoni del calcio. Metodi statistici per l'analisi delle fluttuazioni. Studio delle fisio patologia di malattie monogeniche dello spettro autistico.
23 Maggio 2022 (2h 00m)
Gian Michele Ratto - Corso (attività didattica) - In presenza
Iaging a due fotoni del cloro intracellulare. Modulazione diurna della eccitabilità neuronale.
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