Fixation for Electron Microscopy

Fixation for Electron Microscopy

1st Edition - December 28, 1981
  • Editor: M Hayat
  • eBook ISBN: 9780323147293

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Description

Fixation for Electron Microscopy presents how to better understand the effects of fixatives on the molecular structure of the cell. This book attempts to consider each aspect of fixation, including chemical interactions between fixatives and individual cellular substances. The chemistry of fixative interactions that are discussed in the book is based primarily on the reactions of a fixative with isolated proteins, lipids, nucleic acids, and carbohydrates. The book shows that the correct interpretation of information retrieved from electron micrographs depends on the knowledge of the basic principles underlying the fixation procedure. Also, the book presents the fixation of both eukaryotic and prokaryotic specimens. The special fixation conditions for plant specimens are discussed in detail and have been allotted a whole chapter. Also emphasized in this book is the connection between morphology and biochemical aspects of preparatory treatments and the chemical basis of the formation of artifacts. This topic is useful in understanding the modifications of cell structures introduced during their processing. A guide for recognizing and minimizing major artifacts and fixation faults that are usually encountered is also presented in the book. This valuable resource will prove useful to both students and professionals in the field of biology and clinical medicine. Specimen preservation researchers can also benefit from this book.

Table of Contents


  • Preface

    1 Introduction

    Text

    2 Factors Affecting the Quality of Fixation

    Tissue Specimen Size

    Preparation of Tissue Blocks

    Automatic Specimen Processing

    Osmolarity

    Ionic Composition of Fixative Solution

    Vehicle Osmolality

    Effects of Added Substances

    Methods for Adjusting the Osmolarity

    Recommended Osmolality

    Measurement of Osmolality

    Buffers

    Buffer Types

    Preparation of Buffers

    pH

    Fixative Penetration

    Temperature of Fixation

    Duration of Fixation

    Fixative Concentration

    3 Aldehydes

    Glutaraldehyde (Glutaric Acid Dialdehyde)

    Uses of Glutaraldehyde

    Nature of Commercial Glutaraldehyde

    Reaction with Proteins

    Reaction with Lipids

    Reaction with Nucleic Acids

    Reaction with Carbohydrates

    Effects of Aldehydes on Tissue Physiology

    Osmolality

    Osmolality of Glutaraldehyde

    Temperature

    Concentration

    pH

    Fixation Method with Glutaraldehyde at Higher pHs

    Rate of Penetration

    Shrinkage Caused by Aldehydes

    Heated Glutaraldehyde

    Limitations of Glutaraldehyde

    Purification of Glutaraldehyde

    Determination of Glutaraldehyde Concentration

    Storage of Glutaraldehyde

    Glutaraldehyde-Containing Fixatives

    Formaldehyde

    Reaction with Proteins

    Reaction with Lipids

    Reaction with Nucleic Acids

    Reaction with Carbohydrates

    Preparation of Paraformaldehyde

    Acrolein

    Reaction with Proteins

    Reaction with Lipids

    Precaution in the Handling of Acrolein

    Acrolein Fixatives

    Polyaldehyde

    Role of Aldehydes in Quantitative Electron Microscopy

    Tissue Storage

    4 Osmium Tetroxide

    Introduction

    Reaction with Unsaturated Lipids

    Reaction with Saturated Lipids

    Reaction with Proteins

    Reaction with Lipoproteins

    Reaction with Nucleic Acids

    Reaction with Carbohydrates

    Reaction with Phenolic Compounds

    Loss of Lipids

    Loss of Proteins

    Swelling Caused by Osmium Tetroxide

    Parameters of Fixation

    Concentration of Osmium Tetroxide

    Temperature of Fixation

    Rate of Penetration

    Duration of Fixation

    Osmium Tetroxide as a Primary Fixative

    Osmium Tetroxide as a Vapor Fixative

    Removal of Bound Osmium from Sections

    Osmium Blacks

    Osmeth

    Preparations for and Precautions in the Handling of Osmium

    Tetroxide

    Regeneration of Used Osmium Tetroxide

    5 Permanganates

    Potassium Permanganate Reactions

    Reaction with Membranes

    Reaction with Monoamines

    Reaction with Other Cellular Substances

    Contrast and Loss of Cellular Substances

    Comparison between Potassium Permanganate and Osmium

    Tetroxide

    Fixation Procedures

    Permanganate Fixatives

    6 Miscellaneous Fixatives

    Ruthenium Tetroxide

    Dimethylsuberimidate

    Carbodiimides

    Trioxsalen

    7 Methods of Fixation

    Double Fixation

    Glutaraldehyde and Osmium Tetroxide Mixture

    Simultaneous Fixation for Light and Electron Microscopy

    Anhydrous Fixation

    8 Modes of Fixation

    Vascular Perfusion

    Considerations in Achieving Satisfactory Vascular Perfusion

    Apparatus for Vascular Perfusion

    Methods of Vascular Perfusion

    Immersion Fixation

    Dripping Method

    Injection Method

    Anesthesia

    9 Effects of Fixation

    Effect of Fixation on Actin

    Effect of Fixation on Brain Tissue

    Effect of Fixation on Kidney Tissue

    Renal Medulla

    Biopsy Specimens

    Effect of Fixation on Membrane Fracture

    Effect of Fixation on Mesosomes

    Effect of Fixation on Microtubules and Microfilaments

    Effect of Fixation on Mitochondria

    Effect of Fixation on Myelin

    Effect of Fixation on Plant Virus

    Effect of Fixation on Plasma Membrane

    Effect of Fixation on Staining

    Effect of Fixation on Synaptic Vesicles

    Effect of Fixation on Tight Junctions

    Effect of Fixation on Specimens for X-Ray Microanalysis

    Effect of Fixation on Vascular Endothelium

    Artifacts

    10 Changes in Specimen Volume

    Text

    11 Postmortem Changes

    Text

    12 Plant Specimens

    Text

    13 Fixation for Scanning Electron Microscopy

    Cleaning of Specimens

    Preservation of Microbial Association to Intestinal Epithelium

    Relaxation Procedures

    Fixation Process

    Parameters of Fixation

    Mode of Fixation

    Specimen Size

    Temperature

    Duration of Fixation

    Specimen Shrinkage

    Fixatives

    14 Fixation for Enzyme Cytochemistry

    Comparison of Different Aldehydes

    Effect of Aldehydes on Enzymes

    Factors Affecting the Preservation of Enzyme Activity

    Specimen Size

    Concentration of Aldehyde

    Duration of Fixation

    Temperature

    pH

    Buffers

    Mode of Fixation and Incubation

    Single-Cell Specimens

    Subcellular Fractions

    Cryoultramicrotomy

    15 Fixation for Immunoelectron Microscopy

    Fixatives

    16 Use of Dimethylsulfoxide in Fixation

    Effect on Enzymes

    Effect on Ultrastructure

    17 Criteria for Satisfactory Specimen Preservation

    Text

    18 Artifacts

    Transmission Electron Microscopy

    Scanning Electron Microscopy

    19 Specific Fixation Methods

    Text

    Appendix

    Balanced Salt Solutions

    Commonly Used Salts and Their Physicochemical Properties

    Commonly Used Chemicals

    References

    Index




Product details

  • No. of pages: 521
  • Language: English
  • Copyright: © Academic Press 1981
  • Published: December 28, 1981
  • Imprint: Academic Press
  • eBook ISBN: 9780323147293

About the Editor

M Hayat

Dr. Hayat serves as Distinguished Professor in the Department of Biological Sciences at Kean University. He is an internationally renowned scholar in the fields of electron microscopy and cancer research, has authored and edited more than 31 books, several of these recently with Elsevier (dealing with immunohistochemistry of carcinomas, cancer imaging, and autophagy).

Dr. Hayat has published chapters in 54 books, and authored 10 of them. Seven major publishers have published his books: Cambridge University Press, Elsevier, Academic Press, McGraw-Hill, Plenum Press, CRC Press, and Van Nostrand Reinhold Company. Dr. Hayat’s books have been published in Japanese. He has also published a large number of articles in scientific journals and established the Electron Microscopy Center at Kean University.

Degree Information: Ph.D., Biology, Indiana University

Courses Taught: Electron Microscopy; Cell Biology; Seminar

Primary Area of Expertise: Electron microscopy, cell biology, cancer, health and disease. His research interests include immunohistochemistry and in situ hybridization of human carcinomas, including brain metastases and various primary cancers (lung, breast, prostate, colorectal, pancreatic, ovarian, gastrointestinal, and liver).

Advice For Students Preparing for Your Class: "Read the books and work very hard."

Affiliations and Expertise

Department of Biological Science, Kean University, USA