Note: Front Cover -- Methods in Cell Biology, Volume 30 -- Copyright Page -- Contents -- Preface -- Contributors -- Chapter 1. Image Fidelity: Characterizing the Imaging Tranfer Function -- I. Introduction -- II. The Concept of a Linear, Shift-lnvarient (LSI) System -- III. Characterizing LSI Systems with Sinusoids -- IV. Characterizing the Fluorescence Imaging System -- V. The Contrast Modulation Transfer Function (CMTF) -- VI. Relating the CMTF and the OTF -- VII. Implications -- VIII. The Analysis of the Z-Axis -- IX. Summary -- References -- Chapter 2. Validation of an Imaging System -- I. Shading Correction -- II. Linearity -- III. Geometric Correction and Registration -- IV. Modulation Transfer -- V. Sampling Frequency -- VI. Summary -- References -- Chapter 3. Imaging of Unresolved Objects, Superresolution, and Precision of Distance Measurement, with Video Microscopy -- I. Visualizing Objects Narrower than the Resolution Limit of the Light Microscope -- II. Diffraction Patterns of Very Narrow Objects -- III. Superresolution -- IV. Positional Information and Lateral Setting Accuracy -- V. Ultrathin Optical Sectioning and Axial Setting Accuracy -- VI. Other Factors Affecting the Choice of Pixel Dimensions, Calibra- tion, and Choice of Magnification -- References -- Chapter 4. Fluorescent Standards -- I. Introduction -- II. System Standards -- III. Analytical Calibration Standards -- IV. Concluding Statement -- References -- Chapter 5. Fluorescent Indicators of Ion Concentrations -- I. Introduction -- II. Proton Indicators -- III. Sodium Indicators -- IV. Chloride Indicators -- V. Calcium Indicators -- VI. Prospects and Conclusions -- References -- Chapter 6. Fluorescence Ratio Imaging Microscopy -- I. Introduction -- II. Basic Principles of Fluorescence Ratio Imaging -- III. Requirements for a Fluorescence Ratio Imaging Microscope System. , IV. Validation of the Fluorescence Ratio Imaging Microscope System -- V. Measurements Using Fluorescence Ratio Imaging -- VI. Conclusions and Prospectus -- References -- Chapter 7. Fluorescent Indicators of Membrane Potential: Microspectrofluorometry and Imaging -- I. Introduction -- II. Microspectrofluorometry with Nernstian Dyes -- III. Imaging with Charge-Shift Dyes -- IV. Discussion -- References -- Chapter 8. Resonance Energy Transfer Microscopy -- I. Introduction -- II. Fluorescence Theory -- III. Techniques of Measurement in the Microscope -- IV. Experimental Parameters Measured in the Microscope -- V. Biological Applications -- VI. Problems -- VII. Conclusions -- References -- Chapter 9. Total Internal Reflection Fluorescence Microscopy -- I. Introduction -- II. Theory -- III. Optical Configuration for a Microscope -- IV. Applications of TIRF -- V. Conclusions -- References -- Chapter 10. Designing, Building, and Using a Fluorescence Recovery after Photobleaching Instrument -- 1. Introduction -- II. Designing and Building a Spot FPR Instrument -- III. Artifacts of FPR -- IV. Beyond Spot FPR-Obtaining Spatial Information -- V. Conclusion -- References -- Chapter 11. Interpretation of Fluorescence Correlation Spectroscopy and Photobleaching Recovery in Terms of Molecular Interactions -- I. Introduction -- II. Characteristic Responses -- III. Technical Issues -- IV. Measurement of Aggregation of Fluorophores by FCS -- V. Interaction Systems -- VI. Summary -- References -- Chapter 12. Fluorescence Polarization Microscopy -- I. Introduction -- II. Theory -- III. Experiment -- References -- Chapter 13. Fluorescence Microscopy in Three Dimensions -- I. Introduction -- II. Image Formation in Three Dimensions -- III. Removal of Out-of-Focus Information -- IV. Synthetic Projection and Stereographic Imaging -- V. Experimental Considerations. , VI . Display -- VII. Experimental Results for Imaging Chromosomes -- VIII. Summary -- References -- Chapter 14. Three Dimensional Confocal Fluorescence Microscopy -- I. Introduction -- II. Confocal Microscopy -- III. Confocal Imaging Results -- IV. Discussion -- References -- Chapter 15. Emission of Fluorescence at an Interface -- I. Introduction -- II. Theory -- Ill. Graphical Results -- IV. Consequences for Experiments -- V. Conclusions -- References -- Chapter 16. Fluorescence Labeling and Microscopy of DNA -- I. Introduction -- II. Labeling Strategies -- III. Measurements -- IV. Measurement Techniques -- V. New Directions in FDIM -- References -- Chapter 17. Multiple Spectral Parameter Imaging -- I. Introduction -- II. Practical Aspects of Multiple Spectral Parameter Imaging -- References -- Index -- Content of Resent Volumes.

Note: Front Cover -- Methods in Cell Biology -- Copyright Page -- Contents -- Contributors -- Preface -- Part I: Basic Concept and Preparation Culture Substrates for Cell Mechanical Studies -- Chapter 1: Basic Rheology for Biologists -- I. Introduction and Rationale -- II. Rheological Concepts -- A. Elasticity -- B. Viscosity -- C. Oscillatory Measurements -- III. Rheological Instrumentation -- IV. Experimental Design -- A. Stress-Strain Relation -- B. Stress Relaxation -- C. Creep and Creep Recovery -- D. Frequency Sweep -- E. Time Sweep -- F. Strain or Stress Amplitude Sweeps -- G. Rate-Dependent Viscosity -- H. Flow Oscillation -- V. Sample Preparation -- A. Solids -- B. Liquids and Gelling Systems -- VI. Special Considerations for Biological Samples -- A. Biological Polymers -- B. Intact Tissue -- C. Instrument Selection for Measuring Gelation Kinetics -- VII. Conclusions -- Glossary -- References -- Chapter 2: Polyacrylamide Hydrogels for Cell Mechanics: Steps Toward Optimization and Alternative Uses -- I. Introduction -- II. Principle of the Polyacrylamide Hydrogel -- A. Standard Method of Polymerization -- B. Light-Induced Initiation of Polymerization -- III. Conjugation of Proteins to Polyacrylamide -- A. Carbodiimide-Mediated Cross-Linking -- B. Activation with Acrylic Acid N-Hydroxysuccinimide Ester -- C. Activation with the N-Succinimidyl Ester of Acrylamidohexanoic Acid (N6) -- D. Other Protein-Coupling Methods -- IV. Optimizing the Placement of Beads for Traction Force Microscopy -- V. Manipulation of Gel Geometry -- A. Preparation of Polyacrylamide Microbeads -- B. Preparation of a Model Three-Dimensional Culture System -- VI. Concluding Remarks -- References -- Chapter 3: Microscopic Methods for Measuring the Elasticity of Gel Substrates for Cell Culture: Microspheres, Microindenters, and Atomic Force Microscopy -- I. Introduction. , II. Probing with Microspheres Under Gravitational Forces -- III. Atomic Force Microscopy -- IV. Probing with Spherically Tipped Glass Microindenters -- A. Preparation and Calibration of the Spherically Tipped Microindenter -- B. Calibration of the Microscope and Micromanipulator -- C. Characterization and Calibration of the Microindenter -- D. Measurement of the Indentations of Hydrogels in Response to Forces of the Microindenter -- E. Data Analysis -- F. Discussion -- V. Conclusions -- References -- Chapter 4: Surface Patterning -- I. Introduction -- II. Patterning with Electrodynamic Instabilities -- A. Procedure for Patterning with Electrohydrodynamic Instabilities -- B. Cell Migration on Topographic Surfaces -- III. Lithography Without a Clean Room -- A. Photolithography Basics -- IV. Patterning at the Micro- and Nanoscale with Polymer Mixtures and Block Copolymers -- A. Principles and Procedures for Controlling Pattern Formation with Polymer Mixtures and Block Copolymers -- V. Summary -- Acknowledgments -- References -- Chapter 5: Molecular Engineering of Cellular Environments: Cell Adhesion to Nano-Digital Surfaces -- I. Introduction: Sensing Cellular Environments -- II. Nano-Digital Chemical Surfaces for Regulating Transmembrane-Receptor Clustering -- A. Extended Nanopatterns and Biofunctionalization -- B. Micro-Nanopatterns for Spatially Controlled Molecular Clustering -- C. Cellular Responses to Nano-Digital and Biofunctionalized Surfaces -- D. Local Versus Global Effects of Ligand Density on Cell Adhesion -- E. Cell Spreading and Migration on Different Nanopatterns -- F. High-Resolution Visualization of Cells in Contact with Biofunctionalized Nanopatterns -- III. Outlook for the Future -- Acknowledgments -- References -- Part II: Subcellular Mechanical Properties and Activities. , Chapter 6: Probing Cellular Mechanical Responses to Stimuli Using Ballistic Intracellular Nanorheology -- I. Introduction -- A. Why Cell Mechanics? -- B. Particle-Tracking Nanorheology: Measuring the Local Viscoelastic Properties of a Cell by Tracking the Brownian Motion of Individual Nanoparticles Embedded in the Cell -- C. Local and Global Viscoelastic Properties of the Cell -- D. Interstitial Versus Mesoscale Viscosity of the Cytoplasm -- E. Viscoelastic Properties of the Cytoplasm Depend on the Timescale of Applied Forces -- F. The Viscoelastic Properties of the Cytoplasm Depend on the Amplitude of Applied Forces -- G. Measurements at the Cell Surface Versus in the Cytoplasm -- H. Methods of Delivery of Nanoparticles to the Cytoplasm -- I. BIN: Proof of Principle -- J. Intracellular Micromechanics of Cells in 3D Matrix -- II. Materials and Instrumentation -- A. Preparation of Nanoparticles -- B. Cell Culture -- C. Ballistic Injection of Nanoparticles -- D. Encapsulation of Cells in a 3D Matrix -- E. Imaging of Fluctuating Nanoparticles Embedded in the Cell -- III. Procedures -- A. Preparation of Nanoparticles -- B. Ballistic Injection of Nanoparticles -- C. Cell Seeding and Encapsulation in a 3D Matrix -- D. BIN Analysis -- IV. Pearls and Pitfalls -- V. Concluding Remarks -- A. Unique Advantages of BIN -- B. Advantages of Traditional Particle-Tracking Nanorheology Are Maintained by BIN -- Acknowledgments -- References -- Chapter 7: Multiple-Particle Tracking and Two-Point Microrheology in Cells -- I. Introduction -- II. Principles of Passive Tracer Microrheology -- A. Conventional Passive Microrheology -- B. Expected Tracer Motion and Tracking Performance -- C. Two-Point Microrheology -- III. Multiple-Particle Tracking Algorithms -- A. Image Restoration -- B. Locating Possible Particle Positions. , C. Eliminating Spurious or Unwanted Particle Trajectories -- D. Linking Positions into Trajectories -- E. Available Software Packages and Computing Resources -- IV. Computing Rheology from Tracer Trajectories -- A. Computing Mean-Squared Displacements -- B. Computing Two-Point Mean-Squared Displacements -- C. Applying Automated Image Analysis for Statistics -- D. Converting MSDs to Rheology -- V. Error Sources in Multiple-Particle Tracking -- A. Random Error (Camera Noise) -- B. Systematic Errors -- C. Dynamic Error -- D. Sample Drift, Computational Detrending, and Its Limitations -- E. Effects of Measurement Errors on the MSD -- VI. Instrument Requirements for High-Performance Tracking -- A. Isolation from Vibration, Acoustic Noise, and Thermal Drift -- B. Microscopy-Generation of High-Contrast Tracer Images -- C. Using Low-Noise, Non-interlaced Camera -- D. Using High-Intensity, Filtered Illuminator -- E. Using Synthetic Tracers to Increase Visibility -- VII. Example: Cultured Epithelial Cells -- A. Particle-Tracking Results for TC7 Epithelial Cells -- B. TPM of TC7 Epithelial Cells -- C. Computing Stress Fluctuation Spectra -- VIII. Conclusions and Future Directions -- References -- Chapter 8: Imaging Stress Propagation in the Cytoplasm of a Living Cell -- I. Introduction -- II. Detecting External Stress-Induced Displacements in the Cytoplasm -- A. Green Fluorescent Protein Transfection and Cell Culture -- B. Application of Periodic Mechanical Stress -- C. Image Acquisition -- D. Image Partitioning -- E. Array Shifting -- F. Image Match Searching -- G. Synchronous Displacement -- H. Signal-to-Noise Ratio -- III. Imaging Displacement and Stress Maps in a Live Cell -- A. Quantifying Displacement Maps -- B. Computing Stress Maps from Displacement Maps -- C. Modulation of Stress Distribution Within a Living Cell. , D. Application of Mechanical Loads in Any Direction Using 3D-MTC -- E. Mechanical Anisotropic Signaling to the Cytoskeleton and to the Nucleolus -- F. Limitations of 3D-MTC -- IV. Future Prospects -- Acknowledgments -- References -- Chapter 9: Probing Intracellular Force Distributions by High-Resolution Live Cell Imaging and Inverse Dynamics -- I. Introduction -- II. Methods -- A. Actin Cytoskeleton Mechanics in Cell Protrusion -- B. Force Reconstruction -- C. Probing Heterogeneous Network Elasticity with Speckle Microscopy -- III. Summary -- IV. Appendix -- A. Solution of the Inverse Problem -- Acknowledgments -- References -- Chapter 10: Analysis of Microtubule Curvature -- I. Introduction -- II. Rationale -- III. Raw Data Collection -- A. Point-Click Method -- B. Semiautomated Methods -- C. Data Collection Errors -- IV. Validation Strategy -- A. Modeling of Semiflexible Polymers -- B. Generation of Simulated Data -- C. Validation of Semiflexible Polymer Simulation -- V. Curvature Estimation Methods -- A. Three-Point Method -- B. Shape-Fitting Method -- C. Constructing the Curvature Distribution -- VI. Results -- A. Three-Point Method -- B. Shape-Fitting Method -- VII. Discussion -- VIII. Conclusions -- Acknowledgments -- References -- Chapter 11: Nuclear Mechanics and Methods -- I. Introduction -- II. Experimental Methods for Probing Nuclear Mechanical Properties -- A. Isolation of Individual Nuclei -- B. Micropipette Aspiration Experiments -- C. Substrate Strain Experiments -- D. Compression Experiments -- E. Indentation by AFM -- F. Particle-Tracking Microrheology -- G. Microneedle-Imposed Extension -- III. Discussion and Prospects -- IV. Outlook -- References -- Part III: Cellular and Embryonic Mechanical Properties and Activities -- Chapter 12: The Use of Gelatin Substrates for Traction Force Microscopy in Rapidly Moving Cells. , I. Introduction.

Note: Front Cover -- Methods in Cell Biology, Volume 29 -- Copyright Page -- Contents -- Contributors -- Preface -- Chapter 1. Fluorescent Analog Cytochemistry: Tracing Functional Protein Components in Living Cells -- I. Introduction -- II. Equipping the Laboratory for Fluorescent Analog Cytochemistry -- III. Preparation of Fluorescent Analogs -- IV. Assays of Fluorescent Analogs and Preparations for Microinjection -- V. Delivery of the Conjugates and Handling of Living Cells -- VI. Data Collection and Interpretation -- VII. Prospectus -- References -- Chapter 2. Fluorescent Analogs of Peptides and Hormones -- I. Introduction -- II. Designing a Fluorescent Analog -- III. Characterization of Fluorescent Analogs -- IV. Examples of the Use of Fluorescent Analogs of Hormones and Peptides -- References -- Chapter 3. Fluorescent Analogs of Toxins -- I. Introduction -- II. Examples of Fluorescent Toxins -- III. Selection, Preparation, and Utilization of Fluorescent Toxin Analogs -- IV. Use of Fluorescent Toxin Analogs -- V. Requirements for Successful Application of Toxin Analogs -- References -- Chapter 4. Preparation of Fluorescently Labeled Dextrans and Ficolls -- I. Introduction -- II. Methods -- III. Further Considerations in the Use of Fluorescent Dextrans and Ficolls -- References -- Chapter 5. A Fluorescent Derivative of Ceramide: Physical Properties and Use in Studying the Golgi Apparatus of Animal Cells -- I. Introduction -- II. Synthesis and Purification of C6-NBD-ceramide -- III. Use of Resonance Energy Transfer (RET) to Study C6-NBD-ceramide in Liposomes -- IV. Properties of C6-NBD-ceramide in Cells -- V. Prospectus -- References -- Chapter 6. Fluorescent Labeling of Cell Surfaces -- I. Introduction -- II. Direct Chemical Labels of Cell Surfaces -- III. Fluorescent Ligands as Labels of Cell Surfaces -- References. , Chapter 7. Fluorescent Labeling of Mitochondria -- I. Introduction -- II. Unique Property of Mitochondria: High Membrane Potential -- III. Fluorescent Dyes with Delocalized Positive Charges -- IV. Rhodamine 123 as a Conditional Supravital Dye -- V. Methods of Labeling and Quantitation -- VI. Mitochondrial Localization -- VII. Mitochondrial Morphology -- VIII. Mitochondrial Motility and Distribution -- IX. Monitoring Mitochondrial Membrane Potential -- X. Monitoring Mitochondrial Total Electrochemical Gradient -- XI. Flow Cytometry, Cell Sorting, and Other Applications -- References -- Chapter 8. Fluorescent Labeling of Endoplasmic Reticulum -- I. Introduction -- II. Materials -- III. Labeling Procedures -- IV. Analysis of the Staining -- V. Summary -- References -- Chapter 9. Fluorescent Labeling of Endocytic Compartments -- I. Introduction -- II. Fluorescent Probes -- III. Identifying Compartments of the Endocytic Pathway -- References -- Chapter 10. Incorporation of Macromolecules into Living Cells -- I. Introduction -- II. Strategies for Loading Living Cells -- III. Selected Techniques for Loading Living Cells -- IV. Evaluation and Analysis of Loaded Cells -- References -- Chapter 11. Hapten-Mediated Immunocytochemistry: The Use of Fluorescent and Nonfluorescent Haptens for the Study of Cytoskeletal Dynamics in Living Cells -- I. Introduction -- II. Advantages of Hapten-Mediated Immunocytochemistry -- III. Preparation of Reagents -- IV. Application of Hapten-Mediated Immunocytochemistry -- V. Conclusion and Future Outlook -- VI. Technical Appendix -- References -- Chapter 12. Culturing Cells on the Mircroscope Stage -- I. Introduction -- II. Requirements of the Microscope Culture System -- III. Cell Culture Chambers -- IV. Microscope Incubators -- References -- Chapter 13. Basic Fluorescence Microscopy -- I. Introduction -- II. Microscope Design. , III. Practical Issues of Epiillumination Fluorescence Microscopy -- IV. Basic Setup of an Epiillumination Fluorescence Microscope -- References -- Chapter 14. Quantitative Fluorescence Microscopy Using Photomultiplier Tubes and Imaging Detectors -- I. Introduction -- II. Accessories for Quantitative Fluorescence Analysis -- III. Detectors for Quantitative Fluorescence Microscopy -- References -- Chapter 15. Characteristics of Low Light Level Television Cameras -- I. Introduction -- II. Requirements for a Low Light Level Video Camera -- III. Criteria for Camera Selection and Evaluation -- IV. Performance Characteristics of Existing Low Light Level Cameras -- V. Future Developments -- References -- Chapter 16. Solid-State Imagers for Microscopy -- I. The History of Solid-State Imagers -- II. Theory of CCD Imagers -- III. CCD Camera Implementations -- IV. CCD Performance -- V. Quantitative Imaging with CCDs -- VI. Future Trends in Solid-State Detector Development -- VII. Summary -- References -- Index -- Contents of Recent Volumes.

Note: Literaturangaben , Literaturangaben

Note: Literaturangaben , Literaturangaben