Optical Fluorescence Microscopy From the Spectral to the Nano Dimens
In the last decade, fluorescence microscopy has evolved from a classical “retrospective” microscopy approach into an advanced imaging technique that allows the observation of cellular activities in living cells with increased resolution and dimensions. A
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Alberto Diaspro Editor
Optical Fluorescence Microscopy From the Spectral to the Nano Dimension
Editor Prof. Alberto Diaspro Head, Nanophysics Unit Senior Scientist The Italian Institute of Technology -IIT Via Morego, 30 16163 Genova Italy [email protected] Department of Physics University of Genova Via Dodecaneso, 33 16146 Genova Italy [email protected]
ISBN 978-3-642-15174-3 e-ISBN 978-3-642-15175-0 DOI 10.1007/978-3-642-15175-0 # Springer Heidelberg Dordrecht London New York # Springer-Verlag Berlin Heidelberg 2011 This work is subject to copyright. All rights are reserved, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilm or in any other way, and storage in data banks. Duplication of this publication or parts thereof is permitted only under the provisions of the German Copyright Law of September 9, 1965, in its current version, and permission for use must always be obtained from Springer. Violations are liable to prosecution under the German Copyright Law. The use of general descriptive names, registered names, trademarks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. Cover illustration: High resolution two-photon 3D immunofluorescence image of MAP-2 localised to neauronal-like cells in 11 days old embryoid body; see Fig.13.7 in “Near Infrared 3-Dimensional Nonlinear Optical Monitoring of Stem Cell Differentiation”, Uday. K. Tirlapur and Clarence Yapp Cover design: deblik Berlin, Germany Printed on acid-free paper Springer is part of Springer ScienceþBusiness Media (www.springer.com)
Preface
It was morning, and the new sun sparkled gold across the ripples of a gentle sea. . . . It was another busy day beginning. . . . But way off alone, out by himself beyond boat and shore, Jonathan Livingston Seagull was practising. . .No limits, Jonathan? he thought, and he smiled. His race to learn had begun. – AD free excerption from “Jonathan Livingston Seagull – a story” by Richard Bach-First published in Great Britain by Turnstone Press 1972. This edition published by Element 2003
The intention behind this book is to provide a sort of starting point for those who are interested in the hard task of fully exploiting the capabilities offered by an optical microscopy fluorescence approach. It is fundamental to have some basic concepts about optical microscopy and fluorescence to understand their enormously high potential in basic and applied research. The classical spatial domain of optical microscopy is the submicron level dictated by the physics laws. Notwithstanding this we are moving to the nano-dimension, i.e., a spatial domain from 5 to 100 nm, exploiting the photophysics of the fluorescent markers being used and the technological abilities of detecting low signals and manipulating light in phase and amplitude incl
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