Towards the end of the 1960s, a number of quite different circumstances combined to launch a period of intense activity in the digital processing of electron micro graphs. First, many years of work on correcting the resolution-limiting aberrations of electron microscope objectives had shown that these optical impediments to very high resolution could indeed be overcome, but only at the cost of immense exper imental difficulty; thanks largely to the theoretical work of K. -J. Hanszen and his colleagues and to the experimental work of F. Thon, the notions of transfer func tions were beginning to supplant or complement the concepts of geometrical optics in electron optical thinking; and finally, large fast computers, capable of manipu lating big image matrices in a reasonable time, were widely accessible. Thus the idea that recorded electron microscope images could be improved in some way or rendered more informative by subsequent computer processing gradually gained ground. At first, most effort was concentrated on three-dimensional reconstruction, particu larly of specimens with natural symmetry that could be exploited, and on linear operations on weakly scattering specimens (Chap. l). In 1973, however, R. W. Gerchberg and W. O. Saxton described an iterative algorithm that in principle yielded the phase and amplitude of the electron wave emerging from a strongly scattering speci men.

Preface to Second Edition Several new topics have been added, some small errors have been corrected and some new references have been added in this edition. New topics include aberration corrected instruments, scanning confocal mode of operations, Bloch wave eigenvalue methods and parallel computing techniques. The ?rst edition - cluded a CD with computer programs, which is not included in this edition. - stead the associated programs will be available on an associated web site (currently people.ccmr.cornell.edu/ ̃kirkland,but may move as time goes on). I wish to thank Mick Thomas for preparing the specimen used to record the image in Fig.5.26 and to thank Stephen P. Meisburger for suggesting an interesting biological specimen to use in Fig.7.24. Again, I apologize in advance for leaving out some undoubtedlyoutstanding r- erences. I also apologize for the as yet undiscovered errors that remain in the text. Earl J. Kirkland, December 2009 Preface to First Edition Image simulation has become a common tool in HREM (High Resolution El- tron Microscopy) in recent years. However, the literature on the subject is scattered among many different journals and conference proceedings that have occurred in the last two or three decades. It is dif?cult for beginners to get started in this ?eld.

Computer Techniques for Image Processing in Electron Microscopy: Advances in Electronics and Electron Physics presents the sophisticated computer generated in processing the image. This book discusses the development of fast Fourier transform algorithms, which has led to the possibility of achieving a more reliable interpretation of electron micrographs by digital means. Organized into 10 chapters, this book begins with an overview of image formation in which the properties of the linear approximation are included. This text then reviews the available hardware and the basic mathematical methods of image processing in electron microscopy. Other chapters consider the constraints imposed on the image wave function by the objective lens aperture. This book discusses as well the properties of discrete Fourier transforms. The final chapter deals with a particular processing system called the Improc system. This book is a valuable resource for physicists and researcher workers who are interested in the study of image processing.

The use of computer-based image analysis systems for all kinds of images, but especially for microscope images, has become increasingly widespread in recent years, as computer power has increased and costs have dropped. Software to perform each of the various tasks described in this book exists now, and without doubt additional algorithms to accomplish these same things more efficiently, and to perform new kinds of image processing, feature discrimination and measurement, will continue to be developed. This is likely to be true particularly in the field of three-dimensional imaging, since new microscopy methods are beginning to be used which can produce such data. It is not the intent of this book to train programmers who will assemble their own computer systems and write their own programs. Most users require only the barest of knowledge about how to use the computer, but the greater their understanding of the various image analysis operations which are possible, their advantages and limitations, the greater the likelihood of success in their application. Likewise, the book assumes little in the way of a mathematical background, but the researcher with a secure knowledge of appropriate statistical tests will find it easier to put some of these methods into real use, and have confidence in the results, than one who has less background and experience. Supplementary texts and courses in statistics, microscopy, and specimen preparation are recommended as necessary.

This book has evolved by processes of selection and expansion from its predecessor, Practical Scanning Electron Microscopy (PSEM), published by Plenum Press in 1975. The interaction of the authors with students at the Short Course on Scanning Electron Microscopy and X-Ray Microanalysis held annually at Lehigh University has helped greatly in developing this textbook. The material has been chosen to provide a student with a general introduction to the techniques of scanning electron microscopy and x-ray microanalysis suitable for application in such fields as biology, geology, solid state physics, and materials science. Following the format of PSEM, this book gives the student a basic knowledge of (1) the user-controlled functions of the electron optics of the scanning electron microscope and electron microprobe, (2) the characteristics of electron-beam-sample inter actions, (3) image formation and interpretation, (4) x-ray spectrometry, and (5) quantitative x-ray microanalysis. Each of these topics has been updated and in most cases expanded over the material presented in PSEM in order to give the reader sufficient coverage to understand these topics and apply the information in the laboratory. Throughout the text, we have attempted to emphasize practical aspects of the techniques, describing those instru ment parameters which the microscopist can and must manipulate to obtain optimum information from the specimen. Certain areas in particular have been expanded in response to their increasing importance in the SEM field. Thus energy-dispersive x-ray spectrometry, which has undergone a tremendous surge in growth, is treated in substantial detail.

The book reproduces 55 of more than 300 articles written by the author, representing milestones in methods development of single-particle cryo-EM as well as important results obtained by this technique in the study of biological macromolecules and their interactions. Importantly, neither symmetries nor ordered arrangements (as in two-dimensional crystals, helical assemblies, icosahedral viruses) are required. Although the biological applications are mainly in the area of ribosome structure and function, the elucidation of membrane channel structures and their activation and gating mechanisms are represented, as well. The book is introduced by a commentary that explains the original development of concepts, describes the contributions of the author's colleagues and students, and shows how challenges were overcome as the technique matured. Along the way, the ribosome served as an example for a macromolecule with intricate structure and conformational dynamics that pose challenges for three-dimensional visualization. Toward the end of the book -- bringing us to the present time -- molecular structures with near-atomic resolution are presented, and a novel type of computational analysis, manifold embedding, is introduced. Single-particle cryo-EM is currently revolutionizing structural biology, presenting a powerful alternative to X-ray crystallography as a means to solve the structure of biological macromolecules. The book presents in one place a number of articles containing key advances in mathematical and computational methods leading up to the present time. Secondly, the development of the technique over the years is reflected by ever-expanding discoveries in the field of ribosome structure and function. Thirdly, as all histories of ideas, the history of concepts pertaining to this new method of visualization is fascinating all in itself.

Three-Dimensional Electron Microscopy of Macromolecular Assemblies is the first systematic introduction to single-particle methods of reconstruction. It covers correlation alignment, classification, 3D reconstruction, restoration, and interpretation of the resulting 3D images in macromolecular assemblies. It will be an indispensable resource for newcomers to the field and for all using or adopting these methods.Key Features* Presents methods that offer an alternative to crystallographic techniques for molecules that cannot be crystallized* Describes methods that have been instrumental in exploring the three-dimensional structure of* the nuclear pore complex* the calcium release channel;* the ribosome* chaperonins

This updated and revised edition of a classic work provides a summary of methods for numerical computation of high resolution conventional and scanning transmission electron microscope images. At the limits of resolution, image artifacts due to the instrument and the specimen interaction can complicate image interpretation. Image calculations can help the user to interpret and understand high resolution information in recorded electron micrographs. The book contains expanded sections on aberration correction, including a detailed discussion of higher order (multipole) aberrations and their effect on high resolution imaging, new imaging modes such as ABF (annular bright field), and the latest developments in parallel processing using GPUs (graphic processing units), as well as updated references. Beginning and experienced users at the advanced undergraduate or graduate level will find the book to be a unique and essential guide to the theory and methods of computation in electron microscopy.

Cryoelectron microscopy of biological molecules is among the hottest growth areas in biophysics and structural biology at present, and Frank is arguably the most distinguished practitioner of this art. CryoEM is likely over the next few years to take over much of the structural approaches currently requiring X-ray crystallography, because one can now get good and finely detailed images of single molecules down to as little as 200,000 MW, covering a substantial share of the molecules of greatest biomedical research interest. This book, the successor to an earlier work published in 1996 with Academic Press, is a natural companion work to our forthcoming book on electron crystallography by Robert Glaeser, with contributions by six others, including Frank. A growing number of workers will employ CryoEM for structural studies in their own research, and a large proportion of biomedical researchers will have a growing interest in understanding what the capabilities and limits of this approach are.