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Fundamentals of Light Microscopy and Electronic Imaging, Second Edition provides a coherent introduction to the principles and applications of the integrated optical microscope system, covering both theoretical and practical considerations. It expands and updates discussions of multi-spectral imaging, intensified digital cameras, signal colocalization, and uses of objectives, and offers guidance in the selection of microscopes and electronic cameras, as well as appropriate auxiliary optical systems and fluorescent tags. The book is divided into three sections covering optical principles in diffraction and image formation, basic modes of light microscopy, and components of modern electronic imaging systems and image processing operations. Each chapter introduces relevant theory, followed by descriptions of instrument alignment and image interpretation. This revision includes new chapters on live cell imaging, measurement of protein dynamics, deconvolution microscopy, and interference microscopy. PowerPoint slides of the figures as well as other supplementary materials for instructors are available at a companion website: www.wiley.com/go/murphy/lightmicroscopy
Auteur
DOUGLAS B. MURPHY supervises core facilities in
microscopy and histology at the new HHMI Janelia Farm Research
Campus in Ashburn, Virginia. An Adjunct Professor of Cell Biology
at Johns Hopkins School of Medicine in Baltimore, Maryland, Dr.
Murphy helped establish the School of Medicine Microscope Facility
there, which he supervised until 2006.
MICHAEL W. DAVIDSON is an assistant scholar/scientist
affiliated with the National High Magnetic Field Laboratory and the
Department of Biological Science at Florida State University where
he is involved in developing educational websites. His digital
images and photomicrographs have graced the covers of over 2,000
publications.
Texte du rabat
"This book will provide individuals without background knowledge in optical physics, electronics, or image processing with many of the basic facts they need to know to understand both the power and limitations of their images."
—Cell Biology Education on the First Edition
Fundamentals of Light Microscopy and Electronic Imaging, Second Edition provides a coherent introduction to the principles and applications of the integrated optical microscope system, covering both theoretical and practical considerations. It expands and updates discussions of multi-spectral imaging, intensified digital cameras, signal colocalization, and uses of objectives, and offers guidance in the selection of microscopes and electronic cameras, as well as appropriate auxiliary optical systems and fluorescent tags.
Written in simple, clear language, the book is divided into three sections covering optical principles in diffraction and image formation, basic modes of light microscopy, and components of modern electronic imaging systems and image processing operations. Each chapter introduces relevant theory, followed by descriptions of instrument alignment and image interpretation. Including new sections on live cell imaging, measurement of protein dynamics, deconvolution, multiphoton microscopy, and superresolution microscopy, Fundamentals of Light Microscopy and Electronic Imaging, Second Edition features the following chapters:
Résumé
Fundamentals of Light Microscopy and Electronic Imaging, Second Edition provides a coherent introduction to the principles and applications of the integrated optical microscope system, covering both theoretical and practical considerations. It expands and updates discussions of multi-spectral imaging, intensified digital cameras, signal colocalization, and uses of objectives, and offers guidance in the selection of microscopes and electronic cameras, as well as appropriate auxiliary optical systems and fluorescent tags.
The book is divided into three sections covering optical principles in diffraction and image formation, basic modes of light microscopy, and components of modern electronic imaging systems and image processing operations. Each chapter introduces relevant theory, followed by descriptions of instrument alignment and image interpretation. This revision includes new chapters on live cell imaging, measurement of protein dynamics, deconvolution microscopy, and interference microscopy.
PowerPoint slides of the figures as well as other supplementary materials for instructors are available at a companion website:
www.wiley.com/go/murphy/lightmicroscopy
Contenu
Preface xi
Acknowledgments xii
1. Fundamentals of Light Microscopy 1
Overview 1
Optical Components of the Light Microscope 1
Aperture and Image Planes in a Focused, Adjusted Microscope 5
Note: Objectives, Eyepieces, and Eyepiece Telescopes 6
Koehler Illumination 9
Adjusting the Microscope for Koehler Illumination 9
Note: Summary of Steps for Koehler Illumination 11
Note: Focusing Oil Immersion Objectives 14
Fixed Tube Length versus Infinity Optical Systems 15
Precautions for Handling Optical Equipment 16
Care and Maintenance of the Microscope 17
Exercise: Calibration of Magnification 17
2. Light and Color 21
Overview 21
Light as a Probe of Matter 21
The Dual Particle- and Wave-Like Nature of Light 25
The Quality of Light 26
Properties of Light Perceived by the Eye 27
Physical Basis for Visual Perception and Color 28
Addition and Subtraction Colors 30
Exercise: Complementary Colors 32
3. Illuminators, Filters, and the Isolation of Specific Wavelengths 35
Overview 35
Illuminators and Their Spectra 35
Illuminator Alignment and Bulb Replacement 41
Demonstration: Spectra of Common Light Sources 41
Demonstration: Aligning a 100-W Mercury Arc Lamp in an Epi-Illuminator 43
Filters for Adjusting the Intensity and Wavelength of Illumination 45
Effects of Light on Living Cells 50
4. Lenses and Geometrical Optics 53
Overview 53
Reflection and Refraction of Light 53
Image Formation by a Simple Lens 56
Note: Real and Virtual Images 57
Rules of Ray Tracing for a Simple Lens 58
ObjectImage Math 58
The Principal Aberrations of Lenses 62
Designs and Specifications of Objectives 65
Condensers 71
Oculars 72
Microscope Slides and Coverslips 73
The Care and Cleaning of Optics 73
Exercise: Constructing and Testing an Optical Bench Microscope 76
5. Diffraction and Interference in Image Formation 79
Overview 79
Diffraction and Interference 80
The Diffraction Image of a Point Source of Light 83
The Constancy of Optical Path Length between Object and Image 85
Demonstration: Viewing the Airy Disk with a Pinhole Aperture 85
Effect of Aperture Angle on Diffraction Spot Size 87
Diffraction by a Grating and Calculation of Its Line Spacing, D 89
Demonstration: The Diffraction Grating 93
Abbé's Theory for Image Formation in the Microscope 94
A Diffraction Pattern Is Formed in the Rear Aperture of the Objective 97
Demonstr…