Fluorescence Spectroscopy and Microscopy

Reflecting the expanding field's need for reliable protocols, Fluorescence Spectroscopy and Microscopy: Methods and Protocols offers techniques from a worldwide team of experts on this versatile and vital subject. The topics covered fall into four broad categories: steady-state fluorescence spectroscopy, time-resolved fluorescence spectroscopy, fluorescent probe development, and the various sub-categories of fluorescence microscopy, such as fluorescence recovery after photobleaching (FRAP), live cell FRET imaging (FRETim), fluorescence lifetime imaging (FLIM), fluorescence fluctuation spectroscopy (FFS), and single-molecule fluorescence spectroscopy (smFS). Written as a part of the popular Methods in Molecular Biology series, chapters include the kind of unambiguous detail and key implementation advice that proves essential for successful results.

Comprehensive and practical, Fluorescence Spectroscopy and Microscopy: Methods and Protocols aims to guide both 'novice' and established scientists toward furthering their research with these invaluable techniques.

Explores the methods and applications in the expanding fluorescence field Provides step-by-step detail essential for reproducible results Contains key notes and implementation advice from the experts

Inhalt

Part I: Steady-State Fluorescence Spectroscopy (SSFS). How to Collect National Institute of Standards and Technology (NIST) Traceable Fluorescence Excitation and Emission Spectra.- Steady State Fluorescence Polarization/Anisotropy for the Study of Protein Interactions.- Quantitative Fluorescence Spectral Analysis of Protein Denaturation.- High-Pressure Fluorescence Applications.- Part II: Time-Resolved Fluorescence Spectroscopy (TRFS). Frequency Domain Fluorometry: Theory and Application.- Polar Plot Representation of Time-Resolved Fluorescence.- Ensemble and Single Molecule Detected Time Resolved FRET Methods in Studies of Protein Conformations and Dynamics.- MD + QM Correlations with Tryptophan Fluorescence Spectral Shifts and Lifetimes.- Analysis of Time-Dependent Red Shifts in Fluorescence Emission from Tryptophan Residues in Proteins.- Global Analysis of Time-Resolved Fluorescence Data.- Nanometrology.- Upconversion Spectrophotofluorometry.- Subpicosecond Kerr-Gate Spectrofluorometry.- Photo-Induced Electron Transfer Modeling to Simulate Flavoprotein Fluorescence Decay.- Part III: Fluorescent Probe Development (FPD). Biosynthetic Incorporation of Trp Analogs in Proteins.- Optimization of Fluorescent Proteins.- Monitoring Membrane Properties and Apoptosis Using Membrane Probes of the 3-Hydroxyflavone Family.- Part IV: Fluorescence Microscopy: Fluorescence Recovery After Photobleaching (FRAP). Rectangle FRAP for Measuring Diffusion with a Laser Scanning Microscope.- Part V: Fluorescence Microscopy: Förster Resonance Energy Transfer Imaging (FRETim). A Quantitative Protocol for Intensity-Based Live Cell FRET Imaging.- Part VI: Fluorescence Microscopy: Fluorescence Lifetime Imaging(FLIM). Widefield Fluorescence Lifetime Imaging with Multi-Anode Detectors.- Global Analysis of FRET- FLIM Data in Live Plant Cells.- Time-Resolved Fluorescence Anisotropy Imaging.- Multi-Modal Fluorescence Imaging Spectroscopy.- Part VII: Fluorescence Microscopy: Fluorescence Fluctuation Spectroscopy (FFS). Application of Fluorescence Correlation Spectroscopy (FCS) to Measure the Dynamics of Fluorescent Proteins in Living Cells.- Fluorescence Cross-Correlation Spectroscopy (FCCS) in Living Cells.- Quantifying Lipid-Protein Interaction by Fluorescence Correlation Spectroscopy (FCS).- PET-FCS: Probing Rapid Structural Fluctuations of Proteins and Nucleic Acids by Single-Molecule Fluorescence Quenching.- Z-Scan Fluorescence Correlation Spectroscopy as a Tool for Diffusion Measurements in Planar Lipid Membranes.- Scanning Fluorescence Correlation Spectroscopy (SFCS) with a Scan Path Perpendicular to the Membrane Plane.- Implementation and Application of Pulsed Interleaved Excitation for Dual-Color FCS and RICS.- Quantitative Study of Protein-Protein Interactions in Live Cell by Dual Color Fluorescence Lifetime Cross-Correlation Spectroscopy.- Brightness Experiments.- Global Analysis of Autocorrelation Functions and Photon Counting Distributions in Fluorescence Fluctuation Spectroscopy.- Simulation of Autocorrelation Function and Photon Counting Distribution in Fluorescence Fluctuation Spectroscopy.- Part VIII: Fluorescence Microscopy: Single-Molecule Fluorescence Spectroscopy (smFS). Single-Molecule Fluorescence of Nucleic Acids.- Photoswitchable Fluorescent Proteins for Superresolution Fluorescence Microscopy Above the Diffraction Limit of Light.