This book represents the compilation of papers presented at the second Atomic Force Microscopy/Scanning Tunneling Microscopy (AFM/STM) Symposium, held June 7 to 9, 1994, in Natick, Massachusetts, at Natick Research, Development and Engineering Center, now part ofU.S. Army Soldier Systems Command. As with the 1993 symposium, the 1994 symposium provided a forum where scientists with a common interest in AFM, STM, and other probe microscopies could interact with one another, exchange ideas and explore the possibilities for future collaborations and working relationships. In addition to the scheduled talks and poster sessions, there was an equipment exhibit featuring the newest state-of-the-art AFM/STM microscopes, other probe microscopes, imaging hardware and software, as well as the latest microscope-related and sample preparation accessories. These were all very favorably received by the meeting's attendees. Following opening remarks by Natick's Commander, Colonel Morris E. Price, Jr., and the Technical Director, Dr. Robert W. Lewis, the symposium began with the Keynote Address given by Dr. Michael F. Crommie from Boston University. The agenda was divided into four major sessions. The papers (and posters) presented at the symposium represented a broad spectrum of topics in atomic force microscopy, scanning tunneling microscopy, and other probe microscopies.

Friction force microscopy is an important analytical tool in the field of tribology on the nanometer-scale. The contact area between the probing tip and the sample is reduced to some square nanometers, corresponding to the ideal of a single asperity contact. Traditional concepts, such as friction coefficients, adhesion and elasticity and stick-slip are re-examined with this novel technique. New concepts based upon classical and quantum mechanics are investigated.

Fluorescence microscopy images can be easily integrated into current video and computer image processing systems. People like visual observation; they like to watch a television or computer screen, and fluorescence techniques are thus becoming more and more popular. Since true in vivo experiments are simple to perform, samples can be directly seen and there is always the possibility of manipulating the samples during the experiments; it is an ideal technique for biology and medicine. Images are obtained by a classical (now called wide-field) fluorescence microscope, a confocal scanning microscope, upright or inverted, with epifluorescence or transmission. Computerized image processing may improve definition, and remove glare and scattered light signal. It also makes it possible to compute ratio images (ratio imaging both in excitation and in emission) or lifetime imaging. Image analysis programs may supply a great deal of additional data of various types, starting with calculations of the number of fluorescent objects, their shapes, brightness, etc. Fluorescence microscopy data may be complemented by classical measurement in the cuvette yr by flow cytometry.

Scanning Tunneling Microscopy III provides a unique introduction to the theoretical foundations of scanning tunneling microscopy and related scanning probe methods. The different theoretical concepts developed in the past are outlined, and the implications of the theoretical results for the interpretation of experimental data are discussed in detail. Therefore, this book serves as a most useful guide for experimentalists as well as for theoreticians working in the field of local probe methods.
In this second edition the text has been updated and new methods are discussed.

Biomedical scientists are the foundation of modern healthcare, from cancer screening to diagnosing HIV, from blood transfusion for surgery to food poisoning and infection control. Without biomedical scientists the diagnosis of disease, the evaluation of the effectiveness of treatment, and research into the causes and cures of disease would not be possible. The Fundamentals of Biomedical Science series has been written to reflect the challenges of practicing biomedical science today. It draws together essential basic science with insights into laboratory practice to show how an understanding of the biology of disease is coupled to the analytical approaches that lead to diagnosis. Assuming only a minimum of prior knowledge, the series reviews the full range of disciplines to which a Biomedical Scientist may be exposed-from microbiology to cytopathology to transfusion science. Histopathology describes the processes and practices that are central to the role of the histopathology biomedical scientist, from presampling to diagnosis to laboratory management. It demonstrates throughout how an understanding of cell and tissue physiology is vital to the successful identification of clinical conditions.

This book discusses the various principles in confocal scanning microscopy which has become a useful tool in many practical fields including biological studies and industrial inspection. The methodology presented in this book is unique and is based on the concept of the three-dimensional transfer functions which have been developed by the author and his colleagues over the last five years. With the 3-D transfer functions, resolving power in 3-D confocal imaging can be defined in a unified way, different optical arrangements can be compared with an insight into their inter-relationship, and images of thick objects can be modeled in terms of the Fourier transform which makes the analysis easy. The aim of this book is to provide a systematic introduction to the concept of the 3-D transfer functions in various confocal microscopes, to describe the methods for the derivation of different 3-D transfer functions, and to explain the principles of 3-D confocal imaging in terms of these functions.

Optics at the Nanometer Scale: Imaging and Storing with Photonic Near Fields deals with the fundamentals of and the latest developments and applications of near-field optical microscopy, giving basic accounts of how and under what circumstances superresolution beyond the half- wavelength Rayleigh limit is achieved. Interferometric and fluorescence techniques are also described, leading to molecular and even atomic resolution using light. The storage of optical information at this level of resolution is also addressed.

This volume contains papers presented at the NATO Advanced Research Workshop (ARW) on Photons and Local Probes. The workshop had two predecessors. The first was the NATO ARW on Near Field Optics, held in October 1992 at Arc et Senans and was organized by Daniel Courjon and Dieter Pohl. The other predecessor was a workshop on Photons and Scanning Probe Microscopies held at the University of Konstanz in July 1992. The workshop on Photons and Local Probes was held at the Loechnerhaus on the Reichenau Island at the Lake of Constance, from September 11 to 17, 1994. The Reichenau Island was an important place in Europe in the middle age. Even the tomb of one of the carolingian emperors, Charles the Fat, is located there. At this workshop more than 60 scientists from Europe and the United States met to communicate their latest results in the field of local probes in combination with optical techniques. In eight sessions 31 talks as well as 9 posters were presented. Among those 31 publications were submitted for publication in the NATO proceedings. They were accepted after a strict, but constructive refereeing process.

Arbeitshypothesen sind revidierbar, deklarierten Wahrheiten nicht, sie verkalken zum System; Arbeitshypothesen passen sich den Menschen an, den deklarierten Wahrheiten wird der Mensch angepajJt; die ersten kann mann verwerfen, von den anderen wird man verworfen. FRIEDRICH DORRBNMATI, Nachgedanken Working hypotheses can be revised, ' declared truths cannot-they calcify into dogma. Working hypotheses adapt to people-people adapt to declared truths. One can reject the first but be rejected by the latter. The concept of electron crystallography, i.e., the quantitative use of electron diffraction intensities to solve crystal structures, is by no means new. Based on extensive pioneering efforts on organic and inorganic substances, two major works on electron diffraction structure analysis (or "electronography" as it was then known in Moscow) appeared in English translation during the 19608. These books are B. K. Vainshtein, Strukturnaya Elektronografiya (Structure Analysis by Electron Diffrac tion, translated by E. Feigl andJ. A. Spink, Pergamon Press, Oxford, 1964), and B. B."

Scanning Tunneling Microscopy II, like its predecessor, presents detailed and comprehensive accounts of the basic principles and the broad range of applications of STM and related scanning probe techniques. The applications discussed in this volume come predominantly from the fields of electrochemistry and biology. In contrast to those in STM I, these studies may be performed in air and in liquids. The extensions of the basic technique to map other interactions are described in chapters on scanning force microscopy, magnetic force microscopy, and scanning near-field optical microscopy, together with a survey of other related techniques. Also discussed here is the use of a scanning proximal probe for surface modification. Together, the two volumes give a comprehensive account of experimental aspects of STM and provide essential reading and reference material. In this second edition the text has been updated and new methods are discussed.

The first U. S. Army Natick Research, Development and Engineering Center Atomic Force/Scanning Tunneling Microscopy (AFM/STM) Symposium was held on lune 8-10, 1993 in Natick, Massachusetts. This book represents the compilation of the papers presented at the meeting. The purpose ofthis symposium was to provide a forum where scientists from a number of diverse fields could interact with one another and exchange ideas. The various topics inc1uded application of AFM/STM in material sciences, polymers, physics, biology and biotechnology, along with recent developments inc1uding new probe microscopies and frontiers in this exciting area. The meeting's format was designed to encourage communication between members of the general scientific community and those individuals who are at the cutting edge of AFM, STM and other probe microscopies. It immediately became clear that this conference enabled interdisciplinary interactions among researchers from academia, industry and government, and set the tone for future collaborations. Expert scientists from diverse scientific areas including physics, chemistry, biology, materials science and electronics were invited to participate in the symposium. The agenda of the meeting was divided into three major sessions. In the first session, Biological Nanostructure, topics ranged from AFM ofDNA to STM imagmg ofthe biomoleeule tubulin and bacterialluciferase to the AFM of starch polymer double helices to AFM imaging of food surfaces.

This volume contains the proceedin,r. of the NATO Advanced Study Institute on "Forces in Scanning Probe Methods which was CG-sponsered and organized by the "Forum fUr N anowissenschaften". The conference was held in Schluchsee in the south- em Black Forest (Germany) from March 7-18, 1994. 30 invited lecturers giving tuto- rial talks of historical and recent research activities and about 100 contributed, oral and poster presentations from 130 people participating, created a very active and lestimulating, lively atmosphere. The inventions of scanning tunneling microscopy, atomic force microscopy and near field optical microsocopy opened a new field of research, called scanning probe meth- ods (SPM). During the last decade, the quality of image acquisition made tremendous progress due to advanced data acquisition systems, low noise electronics and suitable mechan- ical and micromechanical constructions. However, a lot of fundamental, unsolved questions about the interaction between probing tip and sample remain. This vol- ume contains 60 contributions dedicated to these problems. Most of the articles are review articles presenting. condensed and relevant information in a way suitable for both students and specialists. Topics that are covered are instrumental aspects, de- signs of force microscopes in various environments, such as ambient pressure, low temperature, ultrahip vacuum and liquids. An important part of the workshop was dedicated to theory, Including all initio calculations and molecular dynamics simula- tions. Mechanical properties, such as adhesion, friction and wear, on the micrometer and nanometer scale were also treated intensively.

Elastic and inelastic scattering in transmission electron microscopy (TEM) are important research subjects. For a long time, I have wished to systematically summarize various dynamic theories associated with quantitative electron micros copy and their applications in simulations of electron diffraction patterns and images. This wish now becomes reality. The aim of this book is to explore the physics in electron diffraction and imaging and related applications for materials characterizations. Particular emphasis is placed on diffraction and imaging of inelastically scattered electrons, which, I believe, have not been discussed exten sively in existing books. This book assumes that readers have some preknowledge of electron microscopy, electron diffraction, and quantum mechanics. I anticipate that this book will be a guide to approaching phenomena observed in electron microscopy from the prospects of diffraction physics. The SI units are employed throughout the book except for angstrom (A), which is used occasionally for convenience. To reduce the number of symbols used, the Fourier transform of a real-space function P'(r), for example, is denoted by the same symbol P'(u) in reciprocal space except that r is replaced by u. Upper and lower limits of an integral in the book are (-co, co) unless otherwise specified. The (-co, co) integral limits are usually omitted in a mathematical expression for simplification. I very much appreciate opportunity of working with Drs. J. M. Cowley and J. C. H. Spence (Arizona State University), J."

Since the first edition of "Scanning 'funneling Microscopy I" has been pub lished, considerable progress has been made in the application of STM to the various classes of materials treated in this volume, most notably in the field of adsorbates and molecular systems. An update of the most recent develop ments will be given in an additional Chapter 9. The editors would like to thank all the contributors who have supplied up dating material, and those who have provided us with suggestions for further improvements. We also thank Springer-Verlag for the decision to publish this second edition in paperback, thereby making this book affordable for an even wider circle of readers. Hamburg, July 1994 R. Wiesendanger Preface to the First Edition Since its invention in 1981 by G. Binnig, H. Rohrer and coworkers at the IBM Zurich Research Laboratory, scanning tunneling microscopy (STM) has devel oped into an invaluable surface analytical technique allowing the investigation of real-space surface structures at the atomic level. The conceptual simplicity of the STM technique is startling: bringing a sharp needle to within a few Angstroms of the surface of a conducting sample and using the tunneling cur rent, which flows on application of a bias voltage, to sense the atomic and elec tronic surface structure with atomic resolution Prior to 1981 considerable scepticism existed as to the practicability of this approach."

Following three printings of the First Edition (1978), the publisher has asked for a Second Edition to bring the contents up to date. In doing so the authors aim to show how the newer microscopies are related to the older types with respect to theoretical resolving power (what you pay for) and resolution (what you get). The book is an introduction to students, technicians, technologists, and scientists in biology, medicine, science, and engineering. It should be useful in academic and industrial research, consulting, and forensics; how ever, the book is not intended to be encyclopedic. The authors are greatly indebted to the College of Textiles of North Carolina State University at Raleigh for support from the administration there for typing, word processing, stationery, mailing, drafting diagrams, and general assistance. We personally thank Joann Fish for word process ing, Teresa M. Langley and Grace Parnell for typing services, Mark Bowen for drawing graphs and diagrams, Chuck Gardner for photographic ser vices, Deepak Bhattavahalli for his work with the proofs, and all the other people who have given us their assistance. The authors wish to acknowledge the many valuable suggestions given by Eugene G. Rochow and the significant editorial contributions made by Elizabeth Cook Rochow."

The main objective of this book is to systematically describe the basic principles of the most widely used techniques for the analysis of physical, structural, and compositional properties of solids with a spatial resolution of approxi mately 1 m or less. Many books and reviews on a wide variety of microanalysis techniques have appeared in recent years, and the purpose of this book is not to replace them. Rather, the motivation for combining the descriptions of various mi croanalysis techniques in one comprehensive volume is the need for a reference source to help identify microanalysis techniques, and their capabilities, for obtaining particular information on solid-state materials. In principle, there are several possible ways to group the various micro analysis techniques. They can be distinguished by the means of excitation, or the emitted species, or whether they are surface or bulk-sensitive techniques, or on the basis of the information obtained. We have chosen to group them according to the means of excitation. Thus, the major parts of the book are: Electron Beam Techniques, Ion Beam Techniques, Photon Beam Techniques, Acoustic Wave Excitation, and Tunneling of Electrons and Scanning Probe Microscopies. We hope that this book will be useful to students (final year undergrad uates and graduates) and researchers, such as physicists, material scientists, electrical engineers, and chemists, working in a wide variety of fields in solid state sciences."

This is the second of three volumes of Methods in Molecular Biology that deal with Physical Methods of Analysis. The first of these, Spectroscopic Methods and Analyses dealt with NMR spec troscopy, mass spectrometry, and metalloprotein techniques, and the third will cover X-ray crystallographic methods. As with the first volume. Microscopy, Optical Spectroscopy, and Macroscopic Techniques is intended to provide a basic understand ing for the biochemist or biologist who needs to collaborate with spe cialists in applying the techniques of modern physical chemistry to biological macromolecules. The methods treated in this book fall into four groups. Part One covers microscopy, which aims to visualize individual molecules or complexes of several molecules. Electron microscopy is the more familiar of these, while scanning tunneling microscopy is a new and rapidly developing tool. Methods for determining the shapes and sizes of molecules in solution are described in Part Two, which includes chapters on X-ray and neutron scattering, light scattering, and ult- centrifugation. Calorimetry, described in Part Three, provides the means to monitor processes involving thermodynamic changes, whether these are intramolecular, such as conformational transition, or the interactions between solutes or between a solute and its sol vent. Part Four is concerned with optical and infrared spectroscopy and describes applications ranging from the measurement of protein concentration by UV absorbance to the analysis of secondary struc ture using circular dichroism and Fourier-transform infrared spec troscopy."

Much of this book was written during a sabbatical visit by J. C. H. S. to the Max Planck Institute in Stuttgart during 1991. We are therefore grateful to Professors M. Ruhle and A. Seeger for acting as hosts during this time, and to the Alexander von Humbolt Foundation for the Senior Scientist Award which made this visit possible. The Ph. D. work of one of us (J. M. Z. ) has also provided much of the background for the book, together with our recent papers with various collaborators. Of these, perhaps the most important stimulus to our work on convergent-beam electron diffraction resulted from a visit to the National Science Foundation's Electron Microscopy Facility at Arizona State University by Professor R. H(lJier in 1988, and from a return visit to Trondheim by J. C. H. S. in 1990. We are therefore particularly grateful to Professor H(lJier and his students and co-workers for their encouragement and collaboration. At ASU, we owe a particular debt of gratitude to Professor M. O'Keeffe for his encouragement. The depth of his under standing of crystal structures and his role as passionate skeptic have frequently been invaluable. Professor John Cowley has also been an invaluable sounding board for ideas, and was responsible for much of the experimental and theoretical work on coherent nanodiffraction. The sections on this topic derive mainly from collaborations by J. C. H. S. with him in the seventies."

This easy-to-follow manual describes tested procedures used to prepare biological samples for scanning and transmission electron microscopy, as well as methods for cytochemistry, immunocytochemistry, and scientific photography. The work is structured to clearly define testing objectives, necessary materials, procedural steps, and expected results; a list of references and trouble shooting techniques round out the text.

Due to its nondestructive imaging power, scanning tunneling microscopy has found major applications in the fields of physics, chemistry, engineering, and materials science. This book provides a comprehensive treatment of scanning tunneling and atomic force microscopy, with full coverage of the imaging mechanism, instrumentation, and sample applications. The work is the first single-author reference on STM and presents much valuable information previously available only as proceedings or collections of review articles. It contains a 32-page section of remarkable STM images, and is organized as a self-contained work, with all mathematical derivations fully detailed. As a source of background material and current data, the book will be an invaluable resource for all scientists, engineers, and technicians using the imaging abilities of STM and AFM. It may also be used as a textbook in senior-year and graduate level STM courses, and as a supplementary text in surface science, solid-state physics, materials science, microscopy, and quantum mechanics.

In 1968, the National Bureau of Standards (NBS) published Special Publication 298 "Quantitative Electron Probe Microanalysis," which contained proceedings of a seminar held on the subject at NBS in the summer of 1967. This publication received wide interest that continued through the years far beyond expectations. The present volume, also the result of a gathering of international experts, in 1988, at NBS (now the National Institute of Standards and Technology, NIST), is intended to fulfill the same purpose. After years of substantial agreement on the procedures of analysis and data evaluation, several sharply differentiated approaches have developed. These are described in this publi cation with all the details required for practical application. Neither the editors nor NIST wish to endorse any single approach. Rather, we hope that their exposition will stimulate the dialogue which is a prerequisite for technical progress. Additionally, it is expected that those active in research in electron probe microanalysis will appreciate more clearly the areas in which further investigations are warranted."

Microcharacterization of materials is a rapidly advancing field. Among the many electron and ion probe techniques, the cathodoluminescence mode of an electron probe instrument has reached a certain maturity, which is reflected by an increas ing number of publications in this field. The rapid rate of progress in applications of cathodoluminescence techniques in characterizing inorganic solids has been especially noticeable in recent years. The main purpose of the book is to outline the applications of cath odoluminescence techniques in the assessment of optical and electronic proper ties of inorganic solids, such as semiconductors, phosphors, ceramics, and min erals. The assessment provides, for example, information on impurity levels derived from cathodoluminescence spectroscopy, analysis of dopant concentra tions at a level that, in some cases, is several orders of magnitude lower than that attainable by x-ray microanalysis, the mapping of defects, and the determination of carrier lifetimes and the charge carrier capture cross sections of impurities. In order to make the book self-contained, some basic concepts of solid-state phys ics, as well as various cathodoluminescence techniques and the processes leading to luminescence phenomena in inorganic solids, are also described. We hope that this book will be useful to both scientists and graduate students interested in microcharacterization of inorganic solids. This book, however, was not intended as a definitive account of cathodoluminescence analysis of in organic solids. In considering the results presented here, readers should re member that many materials have properties that vary widely as a function of preparation conditions."

In 1939, when the electron optics laboratory of Siemens & Halske Inc. began to manufacture the first electron microscopes, the biological and medical profes sions had an unexpected instrument at their disposal which exceeded the reso lution of the light microscope by more than a hundredfold. The immediate and broad application of this new tool was complicated by the overwhelming prob lems inherent in specimen preparation for the investigation of cellular struc tures. The microtechniques applied in light microscopy were no longer appli cable, since even the thinnest paraffin layers could not be penetrated by electrons. Many competent biological and medical research workers expressed their anxiety that objects in high vacuum would be modified due to complete dehydration and the absorbed electron energy would eventually cause degrada tion to rudimentary carbon backbones. It also seemed questionable as to whether it would be possible to prepare thin sections of approximately 0. 5 11m from heterogeneous biological specimens. Thus one was suddenly in posses sion of a completely unique instrument which, when compared with the light microscope, allowed a 10-100-fold higher resolution, yet a suitable preparation methodology was lacking. This sceptical attitude towards the application of electron microscopy in bi ology and medicine was supported simultaneously by the general opinion of colloid chemists, who postulated that in the submicroscopic region of living structures no stable building blocks existed which could be revealed with this apparatus."

vi on geometric probability is included, students can be expected to create a few simple programs like those shown, but for other geometries. I am indebted to Tom Hare for critical reviews of the material and an endless enthusiasm to debate and derive stereological relationships; to John Matzka at Plenum Press for patiently instructing me in the intricacies of typesetting; to Chris Russ for helping to program many of these measurement techniques; and especially to Helen Adams, both for her patience with my creative fever to write yet another book, and for pointing out that the title, which I had intended to contrast to "theoretical stereology," can also be understood as the antonym of "impractical stereology." John C. Russ Raleigh, NC July, 1986 Chapter 1: Statistics 1 Accuracy and precision 1 The mean and standard deviation 5 Distributions 7 Comparison 13 Correlation 18 Nonlinear fitting 19 Chapter 2: Image Types 23 Planar sections 23 Projected images 25 Finite sections 28 Space-filling structures and dispersed phases 29 Types of images and contrast mechanisms 31 Sampling 32 Chapter 3: Manual Methods 35 Volume fraction 35 Surface density 38 Contiguity 41 Mean intercept length 42 Line density 43 Grain size determination 55 Curvature 48 Reticles to aid counting 49 Magnification and units 51 Chapter4: Size Distributions 53 Intercept length in spheres 53 Nonspherical shapes 57 Corrections for finite section thickness 59 Lamellae 61 Measurement of profile size 62 Nonspherical particles 69 vii Contents viii Chapter 5: Computer Metlwds 73