Monday the 9th - Friday the 13th of November, 2009
University of Turku
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Course Director: |
4 ECTS
Target: Starting from the basics of fluorescence the students are familiarized with fluorescence measurement techniques, instrumentation, related chemistry and biochemistry and applications of fluorescence in bioanalytical research.
Tavoite: Muodostaa kokonaiskäsitys fluoresenssin käytön keskeisistä periaatteista ja mahdollisuuksista biolääketieteen ja biologian tutkimuksessa, sekä soveltuvasta instrumentoinnista, laitetekniikasta ja kemiasta.
Contents: Fluorescence basics, Instrumentation and optical components, Spectroscopy, Light detectors and light sources, Labels and label chemistry, Fluorescence applications: FRET, FCS, FRAP, microscopy, cytometry, immunoassays.
Textbook: Alexander P. Demchenko "Introduction to Fluorescence Sensing", Springer
Requirements: Participation on lectures, written essay.
Persons in Charge: Pekka Hänninen, Juhani Soini.
Tentative Schedule:
| Date | Time | Course Program |
| Mo 09.11.2009 Hall: Micro | Lecture 1 | Alexander P. Demchenko: Introduction to fluorescence. Electronic energy levels. Fluorescence and phosphorescence, delayed fluorescence. Display of intermolecular interactions in absorption and emission spectra. Quantum yield. Mechanisms of quenching. |
| Lecture 2 | Alexander P. Demchenko: Fluorescence detection techniques. Intensity-based fluorescence response. Anisotropy-based response and polarization assays. Excimers and exciplexes. Forster resonance energy transfer (FRET) Excited-state electron, charge and proton transfers. Wavelength ratiometry. | |
| Lecture 3 | NN: Fluorescence instrumentation. Spectrophotometers and spectrofluorimeters. Polarization attachments. Lifetime-based instrumentation. Optical components. | |
| Lecture 4 | Alexander P. Demchenko: Fluorescence sensing techniques. Overview of strategies in molecular sensing. Labeled targets in fluorescence assays. Competitor displacement assays. Sandwich assays. Catalytic biosensors. Direct reagent-independent sensing. | |
| Practical work: Demonstration of available spectroscopic instrumentation. | ||
| Tu 10.11.2009 Hall: Salus | Lecture 5 | Alexander P. Demchenko: Quantitative aspects in sensing technologies. Parameters that need to be optimized in every sensor. Determination of binding constants. Modeling the ligand binding isotherm. Kinetics of target binding. Formats for fluorescence detection. |
| Lecture 6 | NN: Analytical applications of fluorescence. Immunodiagnostics. Drug discovery studies. DNA-chips | |
| Lecture 7 | Alexander P. Demchenko: Recognition units: from small organic molecules to biopolymers and cells. Recognition units built from small molecules. Antibodies and their recombinant fragments. Ligand-binding proteins and protein-based display scaffolds. Designed and randomly synthesized peptides. Nucleic acid aptamers. Peptide nucleic acids. Molecularly imprinted polymers. | |
| Lecture 8 | Alexander P. Demchenko: Coupling recognition with fluorescence response. Basic photophysical signal transduction mechanisms. Signal transduction via energy transfer (FRET). Signal transduction via conformational changes. Signal transduction via association and aggregation phenomena. | |
| Lecture 9 | NN: Sensing multiple analytes. Spotted microarrays. Positional encoding. Suspension arrays. Optical encoding. | |
| Practical work: Problem solving in analyte detection. | ||
| We 11.11.2009 Hall: Cal1 | Lecture 10 | Alexander P. Demchenko: Design and properties of fluorescence reporters. Organic dyes and fluorescent proteins The properties of organic dyes. Trp residue as intrinsic reporter in proteins. Dye-doped nanoparticles and dendrimers. Visible fluorescent proteins. Fluorescent conjugated polymers. |
| Lecture 11 | NN or Alexander P. Demchenko: Design and properties of fluorescence reporters. Semiconductor nanocrystals. Semiconductor Quantum Dots and other nanocrystals. Up-converting luminophors. Noble metal nanoparticles and molecular clusters. | |
| Lecture 12 | NN: Reporting based on luminescent metal complexes. Luminescent chelating complexes of lanthanides. Phosphorescent metal complexes. | |
| Lecture 13 | Alexander P. Demchenko: Fluorescence detection of low-molecular targets. Ion detection. Detection of small neutral molecules. High-throughput screening. Optical nose and optical tongue. | |
| Lecture 14 | Alexander P. Demchenko: Fluorescence detection of proteins, nucleic acids and cells. Recognition of protein targets. Nucleic acid detection and sequence identification. Polysaccharides, glycolipids and glycoproteins. Detection of harmful microbes. | |
| Practical work: Problem solving and general discussion. Comparison of properties of different fluorescence reporters. | ||
| Practical work: Demonstration of equipment for multiple analyte analysis. | ||
| Th 12.11.2009 Hall: Cal1 | Lecture 15 | NN: Specialized instrumentation for fluorescence sensing of multiple analytes. Planar waveguides and surface-sensitive detection. Microfluidic devices. Multi-analyte sensor chips and microarrays. Flow cytometers. |
| Lecture 16 | NN: Fluorescence microscopy. Construction of modern fluorescence microscopes. Confocal microscopy. Two- and multiphoton microscopy. Time-resolved microscopy. Molecular recognitions in cell interior. Sensing the whole body (Fluorescence tomography). | |
| Lecture 17 | Alexander P. Demchenko: Fluorescence studies of cell membranes. Structure and dynamic properties of membranes. Membrane potential. Specific biomembrane receptors. | |
| Lecture 18 | NN and Alexander P. Demchenko: Future directions in fluorescence sensing. Sensing on a single molecule level. Genomics, proteomics and other 'omics'. The sensors to any target and to immense number of targets. New level of clinical diagnostics. Advanced sensors in drug discovery. Sensors promising to change the society. | |
| Closing discussion. Instructions for preparation of projects. |
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