Acknowledgements Preface: The Beginning Introduction: The Imaging Chain Foundations, Fundamentals, Principles and Theory Chapter 1 Defining a Science Image A Frame of Reference for the Image in Science The Science Image: a point of departure Criteria for Good Photography Science Photographs require a Scale Photographer's Intent and Subject Matter A picture is worth a thousand words The beginnings of permanent photographs and scientific photography Making the Invisible visible Historical images and Contemporary Point of View Standardized Approaches and Repeatability Father of Standardized Imaging Innovators and technological progress Instrumentation Microscopy and Carl Zeiss The Invisible Spectrum Advancement of Film Technology - Kodak. Agfa, Ilford and Polaroid Short Duration Light, Electric Flash and Stroboscopes Modern Technologies - Digital and Electronic Photography Scanning Electron Microscopy Confocal Microscope Duality of Images Science Images as Art Chapter 2 Human Vision and Perception The Human Visual System The Imaging Room Seeing Basic Structure of the Human Visual System Optics of the Eye and Image Formation Physiology of Seeing Dominant Eye Visual Perception and the Physiological of Sight Perception of color Persistence of Vision Afterimage Perception of Depth Adaption More on Perception Illusions Chapter 3 Applied Physics and Image Formation for the Scientific Photographer Visibility requires Contrast, Magnification, and Resolution Light & Illumination Sources and Spectrums Continuous and Discontinuous Spectrums Color temperature Light behaviours Reflection Refraction Dispersion Interference Lenses Lenses for Scientific Applications Fundamental optics Teleconverters Working Distance Close up Lenses Supplementary Lenses Mirror Lenses Telecentric Lenses Photographic Filters Polarizing Filters Neutral Density Filters Aberrations Curvature of the Field Chromatic Aberrations Depth of Field Diffraction Chapter 4 Digital Cameras, Digital Images, and Strategies The role of the camera Camera Components Shutters Modes of Operation Manual Automatic Secondary Modes of Operation Photographic Exposure Light measurement Types of Shutters Focal Plane Shutters Syncing with electronic flash Electronic shutters Shutter affects on subjects Vibration affects Mirrorless cameras Sensors Pixels Single shot cameras Scanning arrays Multi-shot arrays Sensor sensitivity ISO, Binning, Gain Noise production, dark, shot, sensor and evaluating Sensor evaluation Bit depth Color space Gamma White balance Spectral sensitivity Capture file formats Other file formats Filters Sharpening Color reproduction Noise reduction Digital Artifacts Connecting devices Memory cards Applications, Best Practices and Methods Chapter 5 The Sample and its Role in Laboratory Photography Laboratory Photography Overview The Sample and Treatment Treatment Preparation Selecting a Sample Isolating the sample Objects and characteristics Isolating the subject Composition Handling samples, preparation and treatments Staining and other contrast producing factors Wet samples and immersion methods Making chambers Specimen Tables Surface replicas White, Black or Gray Backgrounds Use of Scales to indicate size Chapter 6 Basic Laboratory Photography Methods: Close-Up Photography, Photomacrography, and Stereomicroscopy Overview of Close-up Photography Close- Up Methods Lenses for Close-Up Photography Supplementary Lenses Extension Tubes Focusing, Depth of Field, and Diffraction Creating Camera to Subject Alignment Selecting the Aperture Exposure in Close-Up applications Photomacrography Introduction Bellows and Laboratory Set-Ups True Macro Lenses and Optical Considerations Other lenses that can be used for magnifications 2:1 and higher Setting up a Macro System Exposure Compensation Exposure Factor equations Depth of Field Stereo Microscopes Photographing with a stereomicroscope Chapter 7 Advanced Laboratory Photography Methods - Making Things Visible Introduction I- Fluorescence Jablonski diagram Ultraviolet and Short Wave Blue Excitation The Fluorescence System II - Photographing with the Invisible spectrum Basic Problems Energy Sources Cameras Lenses Filters Focusing Live View or Auto-focus Exposure Determination Increasing the ISO Noise Reduction Filters Work tethered Multiple Discharges for Electronic Flash Other Strategies III - Polarized light Seeing internal structure The System IV - Schlieren Photographing Schlieren Images V - Scanners as Cameras Scanner Settings Using Descreen Unsharp Masking Imaging Objects on a Scanner VI - Peripheral Photography VII - Stereo and Anaglyphs Making a Stereo Pair Making an Anaglyph VIII - Stroboscopy Chapter 8 A Primer for Lighting Small Laboratory Subjects There is light and then there is lighting Making good light White and Neutral Backgrounds Making Contrast Reducing Contrast Axial lighting Glassware Metal and tent lighting Immersion methods A Working Summary Chapter 9 Light Microscopy I - Foundations and brightfield methods Introduction Fundamentals of Magnified Images Optical Magnification Optical Elements on a Light Microscope Eyepieces Prism Photo or Imaging System Lenses Substage Condendsers Objectives Numerical Aperture Forming Images - Diffraction and Resolution More on Numerical Aperture Objective Corrections Fundamentals useful in Operating a Light Microscope Setting the Eyepieces Focusing Very Small Working Distances Interpupillary Distances Looking into the Body Tube Nosepiece or Turrets Adjusting the Substage Condenser Setting the Field Diaphragm Lamp Setting the Aperture Diaphragm Establishing Proper Brightfield or Kohler Illumination More on Kohler Photographing using a Light Microscope Instrument Cameras DSLR cameras Attaching a Camera to a Microscope II: Advanced Methods Darkfield Differential Interference Contrast Fluorescence Phase Contrast Polarized light Rheinberg Differential Colorization Technique Chapter 10 Confocal Microscopy by James Hayden Introduction Why Confocal ? Types of Confocal Microscopes Fluorescence Microscopy and Confocal Methods Fluorescent Markers Choosing and Working with Fluorophores How a Confocal Microscope Works Balance and Compromises required for forming a Good 2D image Hardware Considerations Lasers Detectors Overview of Instruments Controls and Software Laser Power Detector Settings Simultaneous of Sequential Acquisition Gain and Offset Pinhole Size and Resolution Spatial Resolution and Format Scanning Speed Bidirectional Scanning Digital Zoom Bit depth Averaging / Signal to Noise Accumulation 3D imaging Considerations for Creating an Effective Z stack Consideration for Live Cell Imaging Advanced techniques Chapter 11 Scanning Electron Microscopy by Ted Kinsman Introduction History Modern Machines Theory and Design of Instruments The Nature of an Electron in a Vacuum Electron Source Electron Microscopy Optics Astigmatization The Electron Aperture Resolution in a SEM Signal to Noise Ratio Scan Rotation Specimen Charging Maximizing Resolution Sample Preparation Critical Point Drying Sputter Coating Chapter 12 Ethical Considerations in Scientific Photography: Why Ethics? by James Hayden The Need for Protocols The Image as Data Manipulation and Disclosure Manipulation by Specimen Selection Manipulation by Hardware Settings Manipulation by Imaging Technique Manipulation by Software Manipulation by Presentation Forensic Examination Uncovering Digital Image Fraud Industry Oversight Consequences Conclusions Chapter 13 Considerations and Methods for Image Processing in Science by Staffan Larsson Introduction Terminology: Manipulation, Enhancement, Clarification Software Image J GIMP Adobe Photoshop Basic Color Theory Fundamental Digital Color Models Channels Layers Fundamental Image Editing Methods in Science Monitor Calibration Selection tools and tools overview Image Size Image Editing Tools Overview Selection Image editing tools Pixel Adjustment Tools Image Processing I - Contrast and Color Balance Corrections Method: Setting a white and black point Method: Changing contrast using Levels Method: Using Curves II - Converting RGB files to Grayscale Method: Grayscale Method: Split Channels Method: Channel Mixer Method: Black and White Adjustment Layer III - Sharpening Method: Unsharp Masking Method: High Band Pass Filter Noise reduction using Adobe Camera RAW Method: Eliminating Luminance Noise Method: Despeckle Method: Smart Blur Filter Method: Reducing Noise using the Reduce Noise Filter V - Noise Reduction using the Camera Raw Convertor Software Method: Using the Camera RAW Module VI - Combining fluorescent images VII - Pseudo-coloring B & W images VIII - Making composite images Method: Making a Composite File IX- Type and the Text Tool X - Shapes XI - Preparing files for Publication Method: converting RGB to CMYK Method: Evaluating a CMYK images Black Point Profiles Proofing Gamut Warning Chapter 14 Applications of Computational Photography for Scientist Photographers Image editing and Batch Processing Making actions Increased DOF Making Image Slices Global Image Processing Z Stack file processing using Adobe (R) Photoshop Z Stack file processing using Helicon Focus (R) Z Stack file processing using Zerene Stacker Wide field high resolution Methods Global Image Processing Creating the Image Map High Dynamic Range Images Making Photographic Exposures for HDR Blending the Images Time based imaging Photographic Considerations Intervalometers Making the Photographs Chapter 15 Best Practices Introduction More Thoughts about Best Practices and Workflow The Laboratory and Environmental conditions Cleanliness is imperative Optimizing Camera's Settings Cleaning A Lens Monitors and video displays Color Management Software, upgrades and Optimizing a Computer Image Workflow, Folders, and Naming Files Archiving, Data Redundancy, and Backing Up Planning for Data loss and Disk Failure Digital housekeeping Keeping things Tuned Up Smart phone photography Social Media Conclusion Best Practices Cheat Sheet
Michael Peres is the editor-in-chief of The Focal Encyclopedia of Photography, 4th Edition, and former chair of the biomedical photographic communications department at the Rochester Institute of Technology. Since 1986, he has taught photomicrography, biomedical photography and other applications of photography used in science. Prior to joining the RIT faculty, Peres worked at Henry Ford Hospital and the Charleston Division of West Virginia University as a medical photographer. He is the recipient of the RIT Eisenhart Outstanding Teaching Award and the Schmidt Medal for Lifetime Achievement in the Field of Biocommunications.