Biophysical Tools for Biologists: In Vivo Techniques

Driven in part by the development of genomics, proteomics, and bioinformatics as new disciplines, there has been a tremendous resurgence of interest in physical methods to investigate macromolecular structure and function in the context of living cells. This volume in Methods in Cell Biology is devoted to biophysical techniques in vivo and their applications to cellular biology. Biophysical Tools for Biologists covers methods-oriented chapters on fundamental as well as cutting-edge techniques in molecular and cellular biophysics. This book is directed toward the broad audience of cell biologists, biophysicists, pharmacologists, and molecular biologists who employ classical and modern biophysical technologies or wish to expand their expertise to include such approaches. It will also interest the biomedical and biotechnology communities for biophysical characterization of drug formulations prior to FDA approval.

Describes techniques in the context of important biological problemsDelineates critical steps and potential pitfalls for each method

Section 1. Fluorescence Methods
1) Photoactivation and Photobleaching Techniques for Analysis of Organelle Biogenesis in vivo
2) Analysis of the Dynamics of Living Cells by Fluorescence Correlation Spectroscopy
3) Molecular Sensors Based on Fluorescence Resonance Energy Transfer to Visualize Cellular Dynamics 4) Real-Time Fluorescence of Protein Folding in vivo
5) Microfluidic Glucose Stimulation of Ca+2 Oscillations in Pancreatic Islets

Section 2. Microscopic Methods
6) Introduction to Optical Sectioning: Confocal, Deconvolution, and Two-Photon
7) Use of Electron Tomography to Elucidate Sub-Cellular Structure and Function
8) Proteomics of Macromolecular Complexes by Cellular Cryo-Electron Tomography
9) Total Internal Reflectance Microscopy (TIRF)
10) Atomic Force Microscopy of Living Cells
11) Real-Time Kinetics of Gene Activity in Individual Bacteria
12) Measurement of Cytoskeletal Proteins Globally and Locally in vivo
13) Infrared and Raman Microscopy in Cell Biology
14) Imaging Fluorescent Mice in vivo by Confocal Microscopy
15) Nanoscale Imaging of Intracellular Fluorescent Proteins: Breaking the Diffraction Barrier

Section 3. Methods at the In Vitro/In Vivo Interface
16) Analysis of Protein Posttranslational Modification by Mass Spectrometry
17) Imaging Mass Spectrometry
18) Wet EM Using Quantum Dots
19) Single Cell Capillary Electrophoresis

Section 4. Methods for Diffusion, Viscosity, Force and Displacement
20) Single-Molecule Force Spectroscopy in Living Cells
21) Magnetic Bead Force Applications
22) Measurement of Membrane-Cytoskeleton Adhesion Using Laser Optical Tweezers
23) Cellular Rheological Measurements in vivo
24) Physical Behavior of Cytoskeletal Networks in vitro and in vivo
25) Force Regulation of Microtubule Dynamics in Fission Yeast

Section 5. Techniques for Protein Activity, Protein-Protein and Protein-RNA Interactions
26) Quantifying Protein Activity Using FRET and FLIM Microscopy
27) Measurement of Protein-Protein Interactions in vivo Using FRET and FLIM
28) Measurement of RNA Interactions in vivo Using Molecular Beacons –

Section 6. Computational Modeling
29) Stochastic Modeling in Cell Biology
30) Computational Methods for Analyzing Patterns in Dynamic Biological Phenomena: An Application to Microtubule Dynamics
31) Computational Modeling of Self-Organized Spindle Formation