fret and flim techniques

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FRET and FLIM Techniques
Author : Theodorus W. J. Gadella
Publisher : Elsevier
Release Date : 2011-07-29
ISBN 10 : 9780080915128
Pages : 560 pages

This volume reviews the techniques Förster Resonance Energy Transfer (FRET) and Fluorescence Lifetime Imaging Microscopy (FLIM) providing researchers with step by step protocols and handy hints and tips. Both have become staple techniques in many biological and biophysical fields.

Multiphoton Microscopy and Fluorescence Lifetime Imaging
Author : Karsten König
Publisher : Walter de Gruyter GmbH & Co KG
Release Date : 2018-01-22
ISBN 10 : 311043007X
Pages : 450 pages

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

Methods in Cellular Imaging
Author : Ammasi Periasamy
Publisher : Springer
Release Date : 2013-05-27
ISBN 10 : 1461475139
Pages : 448 pages

Advances in technology have revolutionized the development of light microscopy techniques in biomedical research, thus improving visualization of the microstructure of cells and tissues under physiological conditions. Fluorescence microscopy methods are non-contact and non-invasive and provide high spatial and temporal resolution that other laboratory techniques cannot. This well-illustrated book targets graduate students and scientists who are new to the state-of-the-art fluorescence microscopy techniques used in biological and clinical imaging. It explains basic concepts and imaging procedures for wide-field, confocal, multiphoton excitation, fluorescence resonance energy transfer (FRET), lifetime imaging (FLIM), spectral imaging, fluorescence recovery after photobleaching (FRAP), optical tweezers, total internal reflection, high spatial resolution atomic force microscopy (AFM), and bioluminescence imaging for gene expression. The usage of these techniques in various biological applications, including calcium, pH, membrane potential, mitochondrial signaling, protein-protein interactions under various physiological conditions, and deep tissue imaging, is clearly presented. The authors describe the approaches to selecting epifluorescence microscopy, the detectors, and the image acquisition and processing software for different biological applications. Step-by-step directions on preparing different digital formats for light microscopy images on websites are also provided.

Advanced Fluorescence Microscopy
Author : Peter J. Verveer
Publisher : Humana Press
Release Date : 2016-08-23
ISBN 10 : 9781493942527
Pages : 294 pages

This volume provides an overview of advanced fluorescence microscopy, covering a broad range of methods. Each chapter focuses on a different method and provides a practical guide for application in biological systems. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Advanced Fluorescence Microscopy: Methods and Protocols seeks to provide scientists with methods for biological systems that are of interest.

Molecular Imaging
Author : Ammasi Periasamy,Richard Day
Publisher : Elsevier
Release Date : 2011-04-28
ISBN 10 : 9780080536873
Pages : 312 pages

The detection and measurement of the dynamic interactions of proteins within the living cell are critical to our understanding of cell physiology and pathophysiology. With FRET microscopy and spectroscopy techniques, basic and clinical scientists can make such measurements at very high spatial and temporal resolution. But sources of background information about these tools are very limited, so this book fills an important gap. It covers both the basic concepts and theory behind the various FRET microscopy and spectroscopy techniques, and the practical aspects of using the techniques and analyzing the results. The critical tricks for obtaining a good FRET image and precisely quantitating the signals from living specimens at the nanomolecular level are explained. Valuable information about the preparation of biological samples used for FRET image analysis is also provided. The methods covered include different types of microscopy systems and detectors (wide-field, confocal, multi-photon) as well as specialized techniques such as photobleaching FRET, FLIM-FRET microscopy, spectral imaging FRET, single molecule FRET, and time and image correlation spectroscopy. Starting from the basics, the chapters guide readers through the choice of probes to be used for FRET experiments and the selection of the most suitable experimental approaches to address specific biological questions. Up-to-date, consistently organized, and well-illustrated, this unique book will be welcomed by all researchers who wish to use FRET microscopy and spectroscopy techniques.

Microscopy Techniques
Author : Jens Rietdorf
Publisher : Springer Science & Business Media
Release Date : 2005-06-23
ISBN 10 : 9783540236986
Pages : 319 pages

With contributions by numerous experts

Integrins and Ion Channels
Author : Andrea Becchetti,Annarosa Arcangeli
Publisher : Springer Science & Business Media
Release Date : 2010-12-28
ISBN 10 : 144196066X
Pages : 124 pages

Interdisciplinarity is more often invoked than practised. This is hardly surprising, considering the daunting vastness of modern biology. To reach a satisfactory understanding of a complex biological system, a wide spectrum of conceptual and experimental tools must be applied at different levels, from the molecular to the cellular, tissue and organismic. We believe the multifaceted regulatory interplay between integrin receptors and ion channels offers a rich and challenging field for researchers seeking broad biological perspectives. By mediating cell adhesion to the extracellular matrix, integrins regulate many developmental processes in the widest sense (from cell choice between differentiation and proliferation, to tissue remodeling and organogenesis). Rapidly growing evidence shows that frequent communication takes place between cell adhesion receptors and channel proteins. This may occur through formation of multiprotein membrane complexes that regulate ion fluxes as well as a variety of intracellular signaling pathways. In other cases, cross talk is more indirect and mediated by cellular messengers such as G proteins. These interactions are reciprocal, in that ion channel stimulation often controls integrin activation or expression. From a functional standpoint, studying the interplay between integrin receptors and ion channels clarifies how the extracellular matrix regulates processes as disparate as muscle excitability, synaptic plasticity and lymphocyte activation, just to mention a few. The derangement of these processes has many implications for pathogenesis processes, in particular for tumor invasiveness and some cardiovascular and neurologic diseases. This book provides a general introduction to the problems and methods of this blossoming field.

Advanced Time-Correlated Single Photon Counting Techniques
Author : Wolfgang Becker
Publisher : Springer Science & Business Media
Release Date : 2005-12-20
ISBN 10 : 9783540288824
Pages : 401 pages

In 1984 Desmond O’Connor and David Phillips published their comprehensive book „Time-correlated Single Photon Counting“. At that time time-correlated s- gle photon counting, or TCSPC, was used primarily to record fluorescence decay functions of dye solutions in cuvettes. From the beginning, TCSPC was an am- ingly sensitive and accurate technique with excellent time-resolution. However, acquisition times were relatively slow due to the low repetition rate of the light sources and the limited speed of the electronics of the 70s and early 80s. Moreover, TCSPC was intrinsically one-dimensional, i.e. limited to the recording of the wa- form of a periodic light signal. Even with these limitations, it was a wonderful te- nique. More than 20 years have elapsed, and electronics and laser techniques have made impressive progress. The number of transistors on a single chip has approximately doubled every 18 months, resulting in a more than 1,000-fold increase in compl- ity and speed. The repetition rate and power of pulsed light sources have increased by about the same factor.

FLIM Microscopy in Biology and Medicine
Author : Ammasi Periasamy,Robert M. Clegg
Publisher : CRC Press
Release Date : 2009-07-06
ISBN 10 : 9781420078916
Pages : 472 pages

Detecting Signals at the Single Molecule Level: Pioneering Achievements in Microscopy Recent advances have led to such remarkable improvements in fluorescence lifetime imaging microscopy’s (FLIM) capacity for contrast and sensitivity that researchers can now employ it to detect signals at the single molecule level. FLIM also offers the additional benefit of independence from fluorophore concentration and excitation intensity. Moreover, its unique sensitivity makes it an excellent reporter of conformational changes and of variations in the molecular surroundings of biological molecules. Most of this improvement and discovery have occurred during the past decade, and, to date, information that would benefit a broad range of researchers remains scattered in the literature. Edited by two of the top pioneers in the field, FLIM Microscopy in Biology and Medicine presents the fundamentals of FLIM along with a number of advanced considerations so that a wider audience can appreciate recent and potential improvements that make it such a valuable tool. New Opportunities for Biomedical Researchers... New Challenges for Microscopy Researchers Discussion sections in all the chapters clearly show the challenges for implementing FLIM for various applications. Certain chapters discuss limits on the number of photons required for highly accurate lifetime determinations, as well as the accuracy with which multiple, closely associated lifetime components can reliably be determined. Such considerations are important for the user when he or she is selecting the most advantageous method of FLIM to use for a particular application. While this book provides an introduction for those new to FLIM, it gathers a wealth of material to enhance the work of experts involved in pioneering technological improvements, as well as those research opportunities in this unique and promising area of microscopy.

Time-correlated single photon counting
Author : Desmond O'Connor
Publisher : Academic Press
Release Date : 2012-12-02
ISBN 10 : 0323141447
Pages : 298 pages

Time-correlated Single Photon Counting has been written in the hope that by relating the authors' experiences with a variety of different single photon counting systems, they may provide a useful service to users and potential users of this formidably sensitive technique. Of all the techniques available to obtain information on the rates of depopulation of excited electronic singlet states of molecular species, monitoring of fluorescence provides, in principle, the simplest and most direct measure of concentration. This volume comprises eight chapters, with the first focusing on the time dependence and applications of fluorescence. Succeeding chapters go on to discuss basic principles of the single photon counting lifetime measurement; light sources; photomultipliers; electronics; data analysis; nanosecond time-resolved emission spectroscopy; time dependence of fluorescence anisotropy. This book will be of interest to practitioners in the field of chemistry.

Fluorescence Lifetime Spectroscopy and Imaging
Author : Laura Marcu,Paul M. W. French,Daniel S. Elson
Publisher : CRC Press
Release Date : 2014-07-17
ISBN 10 : 1439861684
Pages : 570 pages

During the past two decades, there has been an increasing appreciation of the significant value that lifetime-based techniques can add to biomedical studies and applications of fluorescence. Bringing together perspectives of different research communities, Fluorescence Lifetime Spectroscopy and Imaging: Principles and Applications in Biomedical Dia

Optical Imaging Techniques in Cell Biology
Author : Guy Cox
Publisher : CRC Press
Release Date : 2006-11-06
ISBN 10 : 1420005618
Pages : 288 pages

Since the word microscopy was coined in 1656, the evolution of the instrument has had a long and convoluted history. Plagued with problems of chromatic aberration, spherical aberration, and challenges with illumination and resolution, the microscope's technical progression happened in a series of fits and starts until the late 19th century. After E