The April 1997 conference held in Prague attracted the cream of primarily European and Russian researchers (with a handful from the US, primarily from the U. of Maryland School of Medicine) to the burgeoning biological and medical applications of innovative optical technology, particularly laser con

This first book on high-speed atomic force microscopy (HS-AFM) is intended for students and biologists who want to use HS-AFM in their research. It provides straightforward explanations of the principle and techniques of AFM and HS-AFM. Numerous examples of HS-AFM studies on proteins demonstrate how to apply this new form of microscopy to specific biological problems. Several precautions for successful imaging and the preparation of cantilever tips and substrate surfaces will greatly benefit first-time users of HS-AFM. In turn, the instrumentation techniques detailed in Chapter 4 can be skipped, but will be useful for engineers and scientists who want to develop the next generation of high-speed scanning probe microscopes for biology. The book is intended to facilitate the first-time use of this new technique, and to inspire students and researchers to tackle their own specific biological problems by directly observing dynamic events occurring in the nanoscopic world. Microscopy in biology has recently entered a new era with the advent of high-speed atomic force microscopy (HS-AFM). Unlike optical microscopy, electron microscopy, and conventional slow AFM, it allows us to directly observe biological molecules in physiological environments. Molecular "movies" created using HS-AFM can directly reveal how molecules behave and operate, without the need for subsequent complex analyses and roundabout interpretations. It also allows us to directly monitor morphological change in live cells, and dynamic molecular events occurring on the surfaces of living bacteria and intracellular organelles. As HS-AFM instruments were recently commercialized, in the near future HS-AFM is expected to become a common tool in biology, and will enhance and accelerate our understanding of biological phenomena.

Since 1995, the noncontact atomic force microscope (NC-AFM) has achieved remarkable progress. Based on nanomechanical methods, the NC-AFM detects the weak attractive force between the tip of a cantilever and a sample surface. This method has the following characteristics: it has true atomic resolution; it can measure atomic force interactions, i.e. it can be used in so-called atomic force spectroscopy (AFS); it can also be used to study insulators; and it can measure mechanical responses such as elastic deformation. This is the first book that deals with all of the emerging NC-AFM issues.

The volumes V, VI and VII will examine the physical and technical foundation for recent progress in applied scanning probe techniques. These volumes constitute a timely comprehensive overview of SPM applications. This is the first book summarizing the state-of-the-art of this technique. The chapters are written by leading researchers and application scientists from all over the world and from various industries to provide a broader perspective.

Scanning tunneling microscopy has achieved remarkable progress and become the key technology for surface science. This book predicts the future development for all of scanning probe microscopy (SPM). Such forecasts may help to determine the course ultimately taken and may accelerate research and development on nanotechnology and nanoscience, as well as all in SPM-related fields in the future.

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."

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 book covers several of the most important topics of current interest at the forefront of scanning probe microscopy. These include a realistic theory of atom-resolving atomic force microscopy (AFM), fundamentals of MBE growth of III-V compound semiconductors and atomic manipulation for future single-electron devices.

Knowledge of the microscopic structure of biological systems is the key to understanding their physiological properties. Most of what we now know about this subject has been generated by techniques that produce images of the materials of interest, one way or another, and there is every reason to believe that the impact of these techniques on the biological sciences will be every bit as important in the future as they are today. Thus the 21st century biologist needs to understand how microscopic imaging techniques work, as it is likely that sooner or later he or she will have to use one or another of them, or will otherwise become dependent on the information that they provide. The objective of this textbook is to introduce its readers to the many techniques now available for imaging biological materials, e.g. crystallography, optical microscopy and electron microscopy, at a level that will enable them to use them effectively to do research. Since all of these experimental methods are best understood in terms of Fourier transformations, this book explains the relevant concepts from this branch of mathematics, and then illustrates their elegance and power by applying them to each of the techniques presented. The book is derived from a one-term course in structural biology that the author gave for many years at Yale. It is intended for students interested either in doing structural research themselves, or in exploiting structural information produced by others. Over the years, the course was taken successfully by advanced undergraduates and by graduate students. Scientists interested in entering the structural biology field later in their careers may also find it useful.

The 14th conference in the series focused on the most recent advances in the study of the structural and electronic properties of semiconducting materials by the application of transmission and scanning electron microscopy. The latest developments in the use of other important microcharacterisation techniques were also covered and included the latest work using scanning probe microscopy and also X-ray topography and diffraction.

The Nobel Prize of 1986 on Sc- ningTunnelingMicroscopysignaled a new era in imaging. The sc- ning probes emerged as a new - strument for imaging with a p- cision suf?cient to delineate single atoms. At ?rstthere were two -the ScanningTunnelingMicroscope,or STM,andtheAtomicForceMic- scope, or AFM. The STM relies on electrons tunneling between tip and sample whereas the AFM depends on the force acting on the tip when itwasplacednearthesample.These were quickly followed by the M- netic Force Microscope, MFM, and the Electrostatic Force Microscope, EFM.TheMFMwillimageasinglemagneticbitwithfeaturesassmallas10nm. WiththeEFMonecanmonitorthechargeofasingleelectron.Prof.PaulHansma atSantaBarbaraopenedthedoorevenwiderwhenhewasabletoimagebiological objects in aqueous environments. At this point the sluice gates were opened and amultitudeofdifferentinstrumentsappeared. There are signi?cant differences between the Scanning Probe Microscopes or SPM, and others such as the Scanning Electron Microscope or SEM. The probe microscopes do not require preparation of the sample and they operate in ambient atmosphere, whereas, the SEM must operate in a vacuum environment and the sample must be cross-sectioned to expose the proper surface. However, the SEM canrecord3Dimage andmovies, featuresthatarenotavailable withthescanning probes. TheNearFieldOpticalMicroscopeorNSOMisalsomemberofthisfamily.At thistimetheinstrumentsuffersfromtwolimitations;1)mostoftheopticalenergy is lost as it traverses the cut-off region of the tapered ?ber and 2) the resolution is insuf?cient for many purposes. We are con?dent that NSOM's with a reasonable opticalthroughputandaresolutionof10nmwillsoonappear.TheSNOMwillthen enterthemainstreamofscanningprobes. VI Foreword In the Harmonic Force Microscope or HFM, the cantilever is driven at the resonantfrequencywiththeamplitudeadjustedsothatthetipimpactsthesampleon each cycle. Theforcesbetween tipandsample generate multiple harmonics inthe motionofthecantilever.Thestrengthoftheseharmonicscanbeusedtocharacterize thephysicalpropertiesofthesurface.

The microanalytical technique of atom probe tomography (APT) permits the spatial coordinates and elemental identities of the individual atoms within a small volume to be determined with near atomic resolution. Therefore, atom probe tomography provides a technique for acquiring atomic resolution three dimensional images of the solute distribution within the microstructures of materials. This monograph is designed to provide researchers and students the necessary information to plan and experimentally conduct an atom probe tomography experiment. The techniques required to visualize and to analyze the resulting three-dimensional data are also described. The monograph is organized into chapters each covering a specific aspect of the technique. The development of this powerful microanalytical technique from the origins offield ion microscopy in 1951, through the first three-dimensional atom probe prototype built in 1986 to today's commercial state-of-the-art three dimensional atom probe is documented in chapter 1. A general introduction to atom probe tomography is also presented in chapter 1. The various methods to fabricate suitable needle-shaped specimens are presented in chapter 2. The procedure to form field ion images of the needle-shaped specimen is described in chapter 3. In addition, the appearance of microstructural features and the information that may be estimated from field ion microscopy are summarized. A brief account of the theoretical basis for processes of field ionization and field evaporation is also included.

Scanning Tunneling Microscopy and its Application presents a unified view of the rapidly growing field of STM,and its many derivatives. A thorough discussion of the various principles provides the background to tunneling phenomena and leads to the many novel scanning-probe techniques, such as AFM, MFM, BEEM, PSTM, etc. After having examined the available instrumentation and the methods for tip and surface preparations, the monograph provides detailed accounts of STM application to metal and semiconductor surfaces, adsorbates and surface chemistry, biology, and nanofabrication. It examines limitations of the present-day investigations and provides hints about possible further trends. This second edition includes important new developments in the field.

Scanning Probe Microscopy - Analytical Methods provides a comprehensive overview of the analytical methods on the nanometer scale based on scanning probe microscopy and spectroscopy. Numerous examples of applications of the chemical contrast mechanism down to the atomic scale in surface physics and chemistry are discussed with extensive references to original work in the recent literature.

Fabrication technologies for nanostructured devices have been developed recently, and the electrical and optical properties of such nanostructures are a subject of advanced research.This book describes the different approaches to spectroscopic microscopy, i.e., Electron Beam Probe Spectroscopy, Spectroscopic Photoelectron Microscopy, and Scanning Probe Spectroscopy. It will be useful as a compact source of reference for the experienced reseracher, taking into account at the same time the needs of postgraduate students and nonspecialist researchers by using a tutorial approach throughout.

The Nobel Prize of 1986 on Sc- ning Tunneling Microscopy sig- led a new era in imaging. The sc- ning probes emerged as a new i- trument for imaging with a pre- sion suf?cient to delineate single atoms. At ?rst there were two - the Scanning Tunneling Microscope, or STM, and the Atomic Force Mic- scope, or AFM. The STM relies on electrons tunneling between tip and sample whereas the AFM depends on the force acting on the tip when it was placed near the sample. These were quickly followed by the - gneticForceMicroscope,MFM,and the Electrostatic Force Microscope, EFM. The MFM will image a single magnetic bit with features as small as 10nm. With the EFM one can monitor the charge of a single electron. Prof. Paul Hansma at Santa Barbara opened the door even wider when he was able to image biological objects in aqueous environments. At this point the sluice gates were opened and a multitude of different instruments appeared. There are signi?cant differences between the Scanning Probe Microscopes or SPM, and others such as the Scanning Electron Microscope or SEM. The probe microscopes do not require preparation of the sample and they operate in ambient atmosphere, whereas, the SEM must operate in a vacuum environment and the sample must be cross-sectioned to expose the proper surface. However, the SEM can record 3D image and movies, features that are not available with the scanning probes.

Introduces both optical microscopy and medical imaging with an emphasis on recurring themes such as resolution and contrast to reinforce understanding. Includes many illustrations and boxed material that give more detailed explanations. Features hands-on activities and experiments. Provides end-of-chapter problems for self-study. Offers supplementary online materials including a solutions manual.

Acoustic microscopy enables the elastic properties of materials to be imaged and measured with the resolution of a good microscope. By using frequencies in the microwave regime, it is possible to make the acoustic wavelength comparable with the wavelength of light, and hence to achieve a resolution comparable with an optical microscope. Solids can support both longitudinal and transverse acoustic waves. At surfaces a unique combination of the two known as Raleigh waves can propagate, and in many circumstances these dominate the contrast in acoustic microscopy. Following the invention of scanning probe microscopes, it is now possible to use an atomic force microscope to detect the acoustic vibration of a surface with resolution in the nanometre range, thus beating the diffraction limit by operating in the extreme near-field. This second edition of Acoustic Microscopy has a major new chapter on the technique and applications of acoustically excited probe microscopy.

Micro-Raman Spectroscopy introduces readers to the theory and application of Raman microscopy. Raman microscopy is used to study the chemical signature of samples with little preperation in a non-destructive manner. An easy to use technique with ever increasing technological advances, Micro-Raman has significant application for researchers in the fields of materials science, medicine, pharmaceuticals, and chemistry.

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."

This monograph focuses on modern femtosecond laser microscopes for two photon imaging and nanoprocessing, on laser tweezers for cell micromanipulation as well as on fluorescence lifetime imaging (FLIM) in Life Sciences. The book starts with an introduction by Dr. Wolfgang Kaiser, pioneer of nonlinear optics and ends with the chapter on clinical multiphoton tomography, the novel high resolution imaging technique. It includes a foreword by the nonlinear microscopy expert Dr. Colin Sheppard. Contents Part I: Basics Brief history of fluorescence lifetime imaging The long journey to the laser and its use for nonlinear optics Advanced TCSPC-FLIM techniques Ultrafast lasers in biophotonics Part II: Modern nonlinear microscopy of live cells STED microscopy: exploring fluorescence lifetime gradients for super-resolution at reduced illumination intensities Principles and applications of temporal-focusing wide-field two-photon microscopy FLIM-FRET microscopy TCSPC FLIM and PLIM for metabolic imaging and oxygen sensing Laser tweezers are sources of two-photon effects Metabolic shifts in cell proliferation and differentiation Femtosecond laser nanoprocessing Cryomultiphoton imaging Part III: Nonlinear tissue imaging Multiphoton Tomography (MPT) Clinical multimodal CARS imaging In vivo multiphoton microscopy of human skin Two-photon microscopy and fluorescence lifetime imaging of the cornea Multiscale correlative imaging of the brain Revealing interaction of dyes and nanomaterials by multiphoton imaging Multiphoton FLIM in cosmetic clinical research Multiphoton microscopy and fluorescence lifetime imaging for resection guidance in malignant glioma surgery Non-invasive single-photon and multi-photon imaging of stem cells and cancer cells in mouse models Bedside assessment of multiphoton tomography

This book highlights important techniques for cellular imaging and covers the basics and applications of electron tomography and related techniques. In addition, it considers practical aspects and broadens the technological focus by incorporating techniques that are only now becoming accessible (e.g. block face imaging). The first part of the book describes the electron microscopy 3D technique available to scientists around the world, allowing them to characterize organelles, cells and tissues. The major emphasis is on new technologies like scanning transmission electron microscopy (STEM) tomography, though the book also reviews some of the more proven technologies like electron tomography. In turn, the second part is dedicated to the reconstruction of data sets, signal improvement and interpretation

This book illustrates the significance of various optical spectroscopy and microscopy techniques, including absorption spectroscopy, fluorescence spectroscopy, infrared spectroscopy, and Raman spectroscopy for deciphering the nature of biological molecules. The content of this book chiefly focuses on (1) the principle, theory, and instrumentation used in different optical spectroscopy techniques, and (2) the application of these techniques in exploring the nature of different biomolecules (e.g., proteins, nucleic acids, enzymes, and carbohydrates). It emphasizes the structural, conformational and dynamic, and kinetic including the changes in biomolecules under a range of conditions. In closing, the book summarizes recent advances in the field of optical spectroscopic and microscopic techniques.

The book is concerned with the theory, background, and practical use of transmission electron microscopes with lens correctors that can correct the effects of spherical aberration. The book also covers a comparison with aberration correction in the TEM and applications of analytical aberration corrected STEM in materials science and biology. This book is essential for microscopists involved in nanoscale and materials microanalysis especially those using scanning transmission electron microscopy, and related analytical techniques such as electron diffraction x-ray spectrometry (EDXS) and electron energy loss spectroscopy (EELS).

This book provides a comprehensive introduction to the methods and variety of Kelvin probe force microscopy, including technical details. It also offers an overview of the recent developments and numerous applications, ranging from semiconductor materials, nanostructures and devices to sub-molecular and atomic scale electrostatics. In the last 25 years, Kelvin probe force microscopy has developed from a specialized technique applied by a few scanning probe microscopy experts into a tool used by numerous research and development groups around the globe. This sequel to the editors' previous volume "Kelvin Probe Force Microscopy: Measuring and Compensating Electrostatic Forces," presents new and complementary topics. It is intended for a broad readership, from undergraduate students to lab technicians and scanning probe microscopy experts who are new to the field.