Low Temperature Biological Microscopy And Microanalysis

The frozen-hydrated specimen is the principal element that unifies the subject of low temperature microscopy, and frozen-hydrated specimens are what this book is all about. Freezing the sample as quickly as possible and then further preparing the specimen for microscopy or microanalysis, whether still embedded in ice or not: there seem to be as many variations on this theme as there are creative scientists with problems of structure and composition to investigate. Yet all share a body of com mon fact and theory upon which their work must be based. Low-Temperature Micros copy and Analysis provides, for the first time, a comprehensive treatment of all the elements to which one needs access. What is the appeal behind the use of frozen-hydrated specimens for biological electron microscopy, and why is it so important that such a book should now have been written? If one cannot observe dynamic events as they are in progress, rapid specimen freezing at least offers the possibility to trap structures, organelles, macro molecules, or ions and other solutes in a form that is identical to what the native structure was like at the moment of trapping. The pursuit of this ideal becomes all the more necessary in electron microscopy because of the enormous increase in resolution that is available with electron-optical instruments, compared to light optical microscopes.

Major improvements in instrumentation and specimen preparation have brought SEM to the fore as a biological imaging technique. Although this imaging technique has undergone tremendous developments, it is still poorly represented in the literature, limited to journal articles and chapters in books. This comprehensive volume is dedicated to the theory and practical applications of FESEM in biological samples. It provides a comprehensive explanation of instrumentation, applications, and protocols, and is intended to teach the reader how to operate such microscopes to obtain the best quality images.

The modem microbiologist is often a real specialist who has difficulty under standing and applying many of the techniques beyond those in his or her own immediate field. On the other hand, most benefits to modem microbiology are obtained when a broad spectrum of scientific approaches can be focused on a problem. In early studies, electron microscopy was pivotal in understanding bacterial and viral morphology, and we still feel that we will understand a disease better if we have seen an electron micrograph of the causative agent. Today, because there is an increased awareness of the need to understand the rela tionships between microbial structure and function, the electron microscope is still one of the most important tools microbiologists can use for detailed analysis of microorganisms. Often, however, the aforementioned modem microbiologist still thinks of ultrastructure as involving negative staining or ultrathin sectioning in order to get a look at the shape of a "bug. " Many of the newer ultrastructure techniques, such as gold-labeled antibody localization, freeze-fracture, X-ray microanalysis, enzyme localization, and even scanning electron microscopy, are poorly under stood by, and therefore forbidding to, the average microbiologist. Even many cell biologists admit to having difficulty staying in touch with current develop ments in the fast-moving field of electron microscopy techniques.

This comprehensive reference illustrates optimal preparation methods in biological electron microscopy compared with common methodological problems. Not only will the basic methodologies of transmission electron microscopy like fixation, microtomy, and microscopy be presented, but the authors also endeavor to illustrate more specialized techniques such as negative staining, autoradiography, cytochemistry, immunoelectron microscopy, and computer-assisted image analysis. Authored by the key leaders in the biological electron microscopy field Illustrates both optimal and suboptimal or artifactual results in a variety of electron microscopy disciplines Introduces students on how to read and interpret electron micrographs