Cell Structure and Function by Microspectrofluorometry

Cell Structure and Function by Microspectrofluorometry

1st Edition - October 28, 1989

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  • Editor: Elli Kohen
  • eBook ISBN: 9781483269733

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Cell Structure and Function by Microspectrofluorometry provides an overview of the state of knowledge in the study of cellular structure and function using microspectrofluorometry. The book is organized into six parts. Part I begins by tracing the origins of modern fluorescence microscopy and fluorescent probes. Part II discusses methods such as microspectroscopy and flow cytometry; the fluorescence spectroscopy of solutions; and the quantitative implementation of fluorescence resonance energy transfer (FRET) in the light microscope. Part III presents studies on metabolism, including the mechanism of action of xenobiotics; biochemical analysis of unpigmented single cells; and cell-to-cell communication in the endocrine and the exocrine pancreas. Part IV focuses on applications of fluorescent probes. Part V deals with cytometry and cell sorting. It includes studies on principles and characteristics of flow cytometry as a method for studying receptor-mediated endocytosis; and flow cytometric measurements of physiologic cell responses. Part VI on bioluminescence discusses approaches to measuring chemiluminescence or bioluminescence in a single cell and measuring light emitted by living cells.

Table of Contents

  • Contributors


    Tomas Hirschfeld—In Memoriam

    Part I History

    1. The Origins of Modern Fluorescence Microscopy and Fluorescent Probes

    I. Introduction

    II. The First Fluorescence Microscopes

    III. Technical Progress

    IV. Advances in Biomedical Applications

    V. Modern Fluorescence Microscopy in Cell and Molecular Biology

    VI. Development of Immunofluorescence


    Part II Methods

    2. Microspectroscopy and Flow Cytometry

    3. From Solution Spectroscopy to Image Spectroscopy

    I. Fluorescence Spectra

    II. Fluorescence Excitation Spectrum

    III. Fluorescence Lifetime and Yield

    IV. Fluorescence Polarization


    4. High-Resolution Fluorescence and Phase Microscopy in Conjunction with Micromanipulation for In Situ Study of Metabolism in Living Cells

    I. Introduction

    II. Resolution of Transmission and Fluorescence Microscopes

    III. Microscope Methods

    IV. Long-Working-Distance Condenser for Micromanipulation

    V. Applications of Spectroscopy to Fluorescence Microscopy

    VI. Instrument Design

    VII. Application of Photography

    VIII. Future Developments


    5. FRET Microscopy: Digital Imaging of Fluorescence Resonance Energy Transfer. Application in Cell Biology

    I. Introduction

    II. Theory of Fluorescence Resonance Energy Transfer

    III. Measurement of FRET: Data Acquisition and Analysis

    IV. Experimental Methods and Results

    V. Discussion and Future Prospects


    6. Fluorescence Scanning Instrumentation

    I. Introduction

    II. Stage Scanning Microfluorometers

    III. Laser Scanning Microfluorometers

    IV. Composition of a Laser Scanning Microscope

    V. Confocal Laser Scanning

    VI. Characteristics of Laser Scanning

    VII. Applications in Laser Scanning


    7. Fluorescence Microscopy in Three Dimensions: Microtomoscopy

    I. Introduction

    II. Confocal Microscopy

    III. Applications

    IV. Discussion


    8. Fluorescence Photochemical Techniques for the Study of Transport in Cytoplasm and Cytoplasmic Models

    I. Introduction

    II. Apparatus and Methodology

    III. Applications to Cytoplasmic Transport

    IV. Applications to Cytoplasmic Models In Vitro

    V. Fluorescence Photoactivation

    VI. Conclusions


    9. Principles of Frequency-Domain Fluorescence Spectroscopy and Applications to Protein Fluorescence

    I. Introduction

    II. Comparison of Time and Frequency-Domain Measurements

    III. Theory of Frequency-Domain Fluorometry

    IV. Tryptophan Fluorescence from Proteins

    V. 2-GHz Frequency-Domain Fluorometry

    VI. Additional Applications of Frequency-Domain Fluorometry

    VII. Future Developments


    10. The First Picosecond in Vision

    I. Introduction

    II. Picosecond Time-Resolved Fluorescence Techniques

    III. Picosecond Fluorescence Spectroscopy Results

    IV. Discussion


    Part III Metabolism

    11. Microspectrofluorometry of Single Living Cells: Quo Vadis

    I. Introduction

    II. Instrumentation and Methods in Microspectrofluorometry

    III. Biological Material

    IV. Spatiotemporal Organization of Cell Metabolism

    V. Spatiotemporal Mapping of Other Organelles: Lysosomes

    VI. Fluorescence Detection of Multiorganelle Complexes Associated with the Cell's Detoxification Function

    VII. Other Applications

    VIII. Conclusions


    12. Mechanism of Action of Xenobiotics: from Molecular Spectral Studies to Microspectrofluorometry of Living Cells

    I. Introduction

    II. Mechanism of Action of Polycyclic Aromatic Hydrocarbons

    III. Mechanism of Action of Antipsoriatic Drugs

    IV. Anticancer Drugs

    V. Conclusions


    13. Microfluorometry as a Tool for Biochemical Analysis in Unpigmented Single Cells

    I. An Example of Convenient Apparatus

    II. Resolution of a Complex Cell Fluorescence Spectrum

    III. Evaluation of Enzymatic Activities in Intact Living Cells

    IV. Use of Fluorescence with Pulsed Excitation

    V. General Conclusions


    14. Fluorescence in the Study of Direct Intercellular Communications: the Case of Pancreatic Cells

    I. Introduction

    II. Fluorescence Approaches to Direct Intercellular Communications

    III. Intercellular Communication Network in the Pancreas

    IV. Concluding Remarks


    Part IV Fluorescent Probes

    15. Approaches to the Study of Spatial and Temporal Changes in the Structure and Chemistry of Cells

    I. Introduction

    II. Approaches to the Study of Cellular Dynamics

    III. Experimental Studies

    IV. Prospectus


    16. Fluorescence Studies of Microtubule Dynamics in Living Cells

    I. Introduction

    II. Microtubule Structure, Intrinsic Polarity, and Organization

    III. Spindle Structure and Function

    IV. Spindle Lability

    V. Fluorescence Approaches to Analyzing Assembly Pathways

    VI. Fluorescence Microscopy, Photobleaching, and Digital Image Processing

    VII. Microtubule Assembly Occurs by a Dynamic Instability Mechanism

    VIII. Comparison with Other Microtubule Arrays

    IX. Future Directions


    17. Optical Measurement of Membrane Potential in Invertebrate Ganglia and Mammalian Cortex

    I. Introduction

    II. Some Optical Signals are Potential-Dependent

    III. Mechanisms

    IV. Dyes

    V. Recording Activity of Individual Neurons in a Molluscan Central Nervous System

    VI. Monitoring Activity in Mammalian Brains

    VII. Summary


    18. Measurement of Free Calcium Concentration inside Single Cells with New Fluorescent Calcium Indicators

    I. Introduction

    II. Methods

    III. Experimental Results

    IV. Applications to Cell Systems

    V. Future Directions


    Part V Cytometry and Cell Sorting

    19. Flow Cytometric Analysis of Ligand Binding and Endocytosis

    I. Introduction

    II. Ligand Binding

    III. Ligand Internalization

    IV. Ligand Acidification

    V. Ligand Degradation

    VI. Conclusion


    20. Flow Cytometric Measurements of Physiologic Cell Responses

    I. Introduction

    II. Physiologic Probes

    III. Instrumentation


    21. Cellular Endogenous Fluorescence: a Basis for Preparing Subpopulations of Functionally Homogeneous Cells

    I. Introduction

    II. Technique of Autofluorescence-Activated Cell Sorting

    III. Purification of Pancreatic B Cells

    IV. Functional Heterogeneity in the Pancreatic B-Cell Population

    V. Subpopulations Homogeneous in Cellular Hormone Content

    VI. Subpopulations Homogeneous in Cellular Glucose Responsiveness

    VII. Subpopulations Homogeneous in Sensitivity to Diabetogenic Agents

    VIII. Conclusions


    Part VI Bioluminescence

    22. Approaches to the Measurement of Chemiluminescence or Bioluminescence in a Single Cell

    I. Introduction

    II. Materials and Methods

    III. Results and Discussion


    23. The Measurement of Light Emitted by Living Cells

    I. Introduction

    II. Electronically Excited States of Molecules

    III. Methods for Absolute Calibration and Measurements in Bioluminescence

    IV. Methods of Detection of Singlet Oxygen in Biological Reactions by Use of Chemiluminescent Probes

    V. The Origin of Bioluminescence

    VI. Bioluminescent Systems

    VII. Colors of Firefly Bioluminescence

    VIII. Experimental Evidence for the Optimization Model of Firefly Fluorescence

    IX. Applications of Firefly Bioluminescence to Environmental Photobiology

    X. Evolution of Bioluminescence in Bacteria

    XI. Emission Spectrum of the Microsomal Chemiluminescence of a Proximate Carcinogen, 7,8-Diol-Benzo(a)Pyrene

    XII. Conclusions



Product details

  • No. of pages: 490
  • Language: English
  • Copyright: © Academic Press 1989
  • Published: October 28, 1989
  • Imprint: Academic Press
  • eBook ISBN: 9781483269733

About the Editor

Elli Kohen

Affiliations and Expertise

University of Miami, Florida, U.S.A.

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