Cell Structure and Function by Microspectrofluorometry - 1st Edition - ISBN: 9780124177604, 9781483269733

Cell Structure and Function by Microspectrofluorometry

1st Edition

Editors: Elli Kohen
eBook ISBN: 9781483269733
Imprint: Academic Press
Published Date: 28th October 1989
Page Count: 490
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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



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




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© Academic Press 1989
Academic Press
eBook ISBN:

About the Editor

Elli Kohen

Affiliations and Expertise

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

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