Fluorescent Biomolecules


Book Description

This volume is based on an international symposium held during September 9-12, 1986 in Bocca di Magra, Italy. The intent of the organizers was to bring together expert practitioners of fluorescence spectroscopy, particularly as applied to biological systems, to assess recent developments in the field and discuss future directions. At the same time the meeting was intended to honor the singular and outstanding scientific career of Gregorio Weber on the occasion of his seventieth birthday. Gregorio Weber is truly the pioneer in the application of fluores cence methods to biochemistry and biophysics. A complete list of his scientific contributions to fluorescence and to protein biochemistry is beyond the scope of this preface. Suffice it to say that since his initial landmark articles on fluorescence, published in the late 1940's and early 1950's, Gregorio Weber has continued to make seminal contribu tions to both the theory and practice of fluorescence and, contrary to many who might be tempted to rest on their laurels, he shows no signs of slackening his pace. In addition to his more obvious tangible contributions to the scientific field, Gregorio Weber has made equally valuable contributions of another type. Specifically, he has had the most profound impact, both profeSSionally and personally, on generations of young scientists.




Fluorescent Proteins


Book Description

This volume brings together cutting-edge laboratory protocols to characterize the novel fluorescent proteins (FPs) and approaches based on fluorescent proteins that aim to answer some of the key cell biological questions. The book covers topics ranging from the database of fluorescent proteins to their characterization and adaptation to a wide range of biological systems. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Fluorescent Proteins: Methods and Protocols serves as an ideal guide for students and academicians enthusiastic about the recent progress in the practical application of fluorescent protein technology.




Fluorescent Methods to Study Biological Membranes


Book Description

Biological membranes play a central role in cell structure, shape and functions. However, investigating the membrane bilayer has proved to be difficult due to its highly dynamic and anisotropic structure, which generates steep gradients at the nanometer scale. Due to the decisive impact of recently developed fluorescence-based techniques, tremendous advances have been made in the last few years in our understanding of membrane characteristics and functions. In this context, the present book illustrates some of these major advances by collecting review articles written by highly respected experts. The book is organized in three parts, the first of which deals with membrane probes and model membranes. The second part describes the use of advanced quantitative and high-resolution techniques to explore the properties of biological membranes, illustrating the key progress made regarding membrane organization, dynamics and interactions. The third part is focused on the investigation of membrane proteins using the same techniques, and notably on the membrane receptors that play a central role in signaling pathways and therapeutic strategies. All chapters provide comprehensive information on membranes and their exploration for beginners in the field and advanced researchers alike.




Molecular Fluorescent Sensors for Cellular Studies


Book Description

Molecular Fluorescent Sensors for Cellular Studies Enables readers to fully understand the fundamentals and chemical principles of fluorescent sensing and the design of fluorescent sensors Fluorescent sensors are able to provide specific chemical information about cells and can be invaluable in understanding processes that underpin health and disease. Molecular Fluorescent Sensors for Cellular Studies provides an avenue into and overview of currently available fluorescent sensing technology and its application to biological imaging. This book aims to help the reader understand the principles of fluorescence and the mechanisms by which fluorescent sensors operate in order to ensure appropriate and optimal use of sensors. Key applications of fluorescent sensing are presented, with explanations not only of how new sensors can be designed, but also how existing sensors can be applied to various biological settings and conditions. Clear and engaging schematics throughout the book explain chemical principles of sensing to the non-expert. Discusses the breadth of fluorescent sensors, from commercially available sensors to those reported in literature which are yet to be used widely Explains how fluorescent sensors operate for appropriate and optimal use from a theoretical standpoint Provides guidance on how to achieve optimal use of fluorescent sensors in practical settings Summarizes the principles behind fluorescent sensors and their design This work will be an invaluable resource for postgraduates and professionals in the fields of microscopy, bioimaging, and diagnostic imaging who wish to harness the information to improve practical applications and to gain key knowledge surrounding the many facets of fluorescent sensing. It is also of interest to medical and biological researchers working across industry, universities and medical institutes.




Fluorescence Spectroscopy in Biology


Book Description

Volume 3 of this new series focuses on brandnew research and applications in biology, biophysics and other fields of life sciences. Many frontline researcher have contributed to this highly attractive and interdisciplinary volume which spans the entire field of present fluorescence spectroscopy including nanotechnology, membrane and DNA studies and fluorescence imaging in cancer research.




Fluorescence Microscopy and Fluorescent Probes


Book Description

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.




Quantitative Biology: Dynamics of Living Systems


Book Description

With the emergence of Systems Biology, there is a greater realization that the whole behavior of a living system may not be simply described as the sum of its elements. To represent a living system using mathematical principles, practical quantities with units are required. Quantities are not only the bridge between mathematical description and biological observations; they often stand as essential elements similar to genome information in genetics. This important realization has greatly rejuvenated research in the area of Quantitative Biology. Because of the increased need for precise quantification, a new era of technological development has opened. For example, spatio-temporal high-resolution imaging enables us to track single molecule behavior in vivo. Clever artificial control of experimental conditions and molecular structures has expanded the variety of quantities that can be directly measured. In addition, improved computational power and novel algorithms for analyzing theoretical models have made it possible to investigate complex biological phenomena. This research topic is organized on two aspects of technological advances which are the backbone of Quantitative Biology: (i) visualization of biomolecules, their dynamics and function, and (ii) generic technologies of model optimization and numeric integration. We have also included articles highlighting the need for new quantitative approaches to solve some of the long-standing cell biology questions. In the first section on visualizing biomolecules, four cutting-edge techniques are presented. Ichimura et al. provide a review of quantum dots including their basic characteristics and their applications (for example, single particle tracking). Horisawa discusses a quick and stable labeling technique using click chemistry with distinct advantages compared to fluorescent protein tags. The relatively small physical size, stability of covalent bond and simple metabolic labeling procedures in living cells provides this type of technology a potential to allow long-term imaging with least interference to protein function. Obien et al. review strategies to control microelectrodes for detecting neuronal activity and discuss techniques for higher resolution and quality of recordings using monolithic integration with on-chip circuitry. Finally, the original research article by Amariei et al. describes the oscillatory behavior of metabolites in bacteria. They describe a new method to visualize the periodic dynamics of metabolites in large scale cultures populations. These four articles contribute to the development of quantitative methods visualizing diverse targets: proteins, electrical signals and metabolites. In the second section of the topic, we have included articles on the development of computational tools to fully harness the potential of quantitative measurements through either calculation based on specific model or validation of the model itself. Kimura et al. introduce optimization procedures to search for parameters in a quantitative model that can reproduce experimental data. They present four examples: transcriptional regulation, bacterial chemotaxis, morphogenesis of tissues and organs, and cell cycle regulation. The original research article by Sumiyoshi et al. presents a general methodology to accelerate stochastic simulation efforts. They introduce a method to achieve 130 times faster computation of stochastic models by applying GPGPU. The strength of such accelerated numerical calculation are sometimes underestimated in biology; faster simulation enables multiple runs and in turn improved accuracy of numerical calculation which may change the final conclusion of modeling study. This also highlights the need to carefully assess simulation results and estimations using computational tools.




Understanding Light Microscopy


Book Description

Introduces readers to the enlightening world of the modern light microscope There have been rapid advances in science and technology over the last decade, and the light microscope, together with the information that it gives about the image, has changed too. Yet the fundamental principles of setting up and using a microscope rests upon unchanging physical principles that have been understood for years. This informative, practical, full-colour guide fills the gap between specialised edited texts on detailed research topics, and introductory books, which concentrate on an optical approach to the light microscope. It also provides comprehensive coverage of confocal microscopy, which has revolutionised light microscopy over the last few decades. Written to help the reader understand, set up, and use the often very expensive and complex modern research light microscope properly, Understanding Light Microscopy keeps mathematical formulae to a minimum—containing and explaining them within boxes in the text. Chapters provide in-depth coverage of basic microscope optics and design; ergonomics; illumination; diffraction and image formation; reflected-light, polarised-light, and fluorescence microscopy; deconvolution; TIRF microscopy; FRAP & FRET; super-resolution techniques; biological and materials specimen preparation; and more. Gives a didactic introduction to the light microscope Encourages readers to use advanced fluorescence and confocal microscopes within a research institute or core microscopy facility Features full-colour illustrations and workable practical protocols Understanding Light Microscopy is intended for any scientist who wishes to understand and use a modern light microscope. It is also ideal as supporting material for a formal taught course, or for individual students to learn the key aspects of light microscopy through their own study.




Three-Dimensional Confocal Microscopy: Volume Investigation of Biological Specimens


Book Description

The integration of confocal microscopy and volume investigation has led to an unprecedented ability to examine spatial relationships between cellular structure and function. The goal of this book is to familiarize the reader with these new technologies and to demonstrate their applicability to a wide range of biological and clinical problems. - Volume investigation - Three-dimensional reconstruction - Fluroescent probe design - Biological applications of confocal microscopy, including calcium imaging, receptor movement, and diagnostic pathology - Confocal data display and analysis - Twenty-eight pages of color




Advanced Fluorescence Reporters in Chemistry and Biology I


Book Description

Fluorescence reporter is the key element of any sensing or imaging technology. Its optimal choice and implementation is very important for increasing the sensitivity, precision, multiplexing power, and also the spectral, temporal, and spatial reso- tion in different methods of research and practical analysis. Therefore, design of ?uorescence reporters with advanced properties is one of the most important problems. In this volume, top experts in this ?eld provide advanced knowledge on the design and properties of ?uorescent dyes. Organic dyes were the ?rst ?uorescent materials used for analytical purposes, and we observe that they retain their leading positions against strong competition of new materials – conjugated polymers, semiconductor nanocrystals, and metal chelating complexes. Recently, molecular and cellular biology got a valuable tool of organic ?uorophores synt- sized by cell machinery and incorporated into green ?uorescent protein and its analogs. Demands of various ?uorescence techniques operating in spectral, anisotropy, and time domains require focused design of ?uorescence reporters well adapted to these techniques. Near-IR spectral range becomes more and more attractive for various applications, and new dyes emitting in this range are strongly requested. Two-photonic ?uorescence has become one of the major tools in bioimaging, and ?uorescence reporters well adapted to this technique are in urgent need. These problems cannot be solved without the knowledge of fundamental principles of dye design and of physical phenomena behind their ?uorescence response.