Image Acquisition

To achieve the best analysis results, you must start with a quality image. Image-Pro offers a streamlined image acquisition dialogue that makes it easy to acquire good images. It offers the necessary tools for basic 2D imaging as well as those needed for sophisticated multi-dimensional experiments. With Image-Pro Plus, you can acquire files directly from cameras, scanners, disks or CD-ROM. Visit the Supported Devices section to see which devices are supported by Image-Pro.

Automated Microscope Control
Control complex microscopy equipment in a simple, repeatable manner for reproducible results. Automatically control your microscope stage in any X, Y, or Z direction and eliminate the need to manually adjust microscope settings.

Multichannel Acquisition

Automate and manage all combinations of image acquisition modes and image sets including time, channel (wavelength), focus (z-stack) and stage position.

Composite image derived from16 Z-stack sets from 3 fluorescent channels - Image courtesy of Richard Cole, Wadsworth Center, Albany, NY

• Configuration Wizard
  The Configuration Wizard guides you through the necessary setup procedures to ensure the success of your acquisition. Image-Pro will automatically check to make sure it has the necessary settings to acquire your images, including objective lenses, spatial calibrations, camera settings, and ookup table information.

• Exposure Control
  Full exposure control is available for each channel. You may even specify a composite image to be automatically built as the image set is acquired. Convenient testing options ensure that no time is wasted in method development and an optimum image set is produced. For best results, background correction may be implemented.
• Status Update
  Monitor the progress of your acquisition with the status display. View the total elapsed time, time left between cycles, current sample position, wavelength, and Z-position. You may also pause your acquisition and re-focus at the current position. If you pause in the middle of a Z-stack, the partial Z-stack will be discarded and a new stack, centered on the new focus position will be acquired.

Time Lapse Acquisition
Investigate changes in live specimens over time by acquiring images at predefined intervals. Play your time lapse images as a movie to view movement and other activities.

Live Analysis & Measurements
Analyze samples live, without the need to acquire. The Workspace Preview feature makes it easy to count objects, apply measurements and interact with online images. The optional Live Tiling and Live Extended Depth of Field tools make it easy to tile large images and perform EDF operations live - while acquiring your images.

Brain cortex cell image captured using Live Tiling - Image courtesy of Joop van Heerikhuize,
Netherlands Institute for Neuroscience

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Live Tiling & Live EDF

Image-Pro Plus 7.0 offers the optional Live Tiling and Live Extended Depth of Field (EDF) module, which makes it easy to acquire large, tiled images and beautifully focused images – live – without an automated stage or focus drive. Minimizing the number of steps required to tile images or created focused images, Live Tiling and Live EDF allows you to acquire the images you need, within minutes.

Live Tiling
Live Tiling Live Tiling lets you expand the field of your captured image, using a manual X-Y stage. This tool makes it easy for you to tile and stitch adjacent fields – live – without an automated stage. Simply move your stage and watch the image field of view grow until you have covered your entire specimen. All you need is a microscope, manual X-Y stage and a camera with Image-Pro drivers.

Live Tiling Details:

• No motor stage or focus drive needed
• Works with any Image-Pro Color or Mono camera/driver
• Obtains a much higher resolution image than using a low mag objective

Live EDF
Live EDF allows you to capture a fully-focused image for specimens whose thickness exceeds the microscope depth of field. Available for any manual microscope, Live EDF solves the depth of field issues that often plague Entomologist, Botanists, and Parts and Material manufacturers.

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Process & Enhance

Use sophisticated image processing and enhancement tools to uncover details within your images.

Imaging Tiling
Tile together a series of images or align sequence images to see a full representation of your research sample.

Before Tiling	After Tiling

Image Alignment
Automatically align either individual or sequence images.

Extended Depth of Field
Create a focused image from a series of unfocused images with Extended Depth of Field.

Filter and Enhance
Choose from a variety of enhancement and edge filters to filter noise and improve image detail.

• Work with Filter dialogs that include descriptions and interactive preview windows
• Use Enhancement filters that include Low-Pass, Hi-Pass, Gaussian, Hi-Gaussian, Sharpen, Flatten, Median,
• Rank, Local Equalization, and De-speckle in user-definable kernel sizes
• Outline objects with various Edge Filter methods including Sobel, Roberts, Laplace, Variance, Phase, Horizontal, and Vertical
• Process objects with Morphological filters including Erode, Dilate, Open, Close, Top Hat, Well, Branch/End,
• Watershed, Thinning, Pruning, Distance, and Reduce in user-definable kernel sizes
• Work with Large Spectral Filters: Fast-acting, large kernel size (up to 4000 x 4000) filters that include Low-Pass, Hi-Pass, Band-Pass, and Edge filters
• Apply filters to individual frames or to entire sequence
• Customize kernel files for morphological and convolution filters

Original image	Sobel edge filter applied

Morphology
Use a wide range of morphological filters to precisely segment structures and prepare images for automatic measurement.

Background Subtraction & Correction
Eliminate background noise and correct uneven backgrounds in your images for improved analysis.

Original image	Background subtracted	New image

Merge Color Channels
Use the Color Composite feature to merge multiple fluorescent images acquired as monochrome single wavelengths into a color composite image.

Composite of tinted gray scale images

Co-localization
Detect co-localization in biological specimens and graphically display the association between the two sets of data in a scatter plot.

Detect co-localization in fluorescent images - Mouse fibroblast cells - microtubules stained with FITC, mitochondria stained with Texas Red, nuclei stained with DAPI.

Pseudo-color
Use pseudo-color to highlight features of interest in a gray scale image. Use colors to visually amplify specific intensities which are normally difficult to distinguish from their surroundings.

Fast Fourier Transforms
Perform forward and inverse Fast Fourier Transforms (FFTs) to eliminate artifacts.

• Edit the spectrum directly including Low-Pass, High-Pass, un-sharpen, spike cut, and spike boost
• Save and reload data

Before FFT	After FFT

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Measure & Classify

Count and Classify Objects
Count and Characterize objects using over 50 manual and automatic measurement tools. Tag objects of interest and easily sort by size or other measurement parameters.

• Measure lengths, roundness, aspects, angles, areas, perimeters, and holes
• Set ranges for measurements to filter unwanted objects
• Group objects in classes based on measurements
• Resolve clustered objects with Watershed, Auto-Split, and Cluster analysis tools
• Use colors to separate and measure objects in color images
• Export measurements to statistical and spreadsheet package via DDE
• Display measurements as histograms & scattergrams

Count and classify objects - Image courtesy of Dr. Gabriel Corkidi Blanco, Centro de Investigacion Aplicada y Desarollo Technologico, UNAM, Morelo, Mexico

Threshold and measure the percent area of objects.

Threshold & Segmentation Tools
Use threshold and segmentation tools to extract objects of interest and distinguish them from other objects or from the background image.

Original image	Threshold values are set
Image after setting threshold	Mask image based on threshold values

Calibration Tools
Use the Image-Pro Plus calibration tools to redefine the scale of an image to express values in other units of measure. Learn more about Image-Pro Plus calibrations in the Spatial Calibration Product Note:

• Calibration wizard helps you to calibrate an active image and create reference and system calibrations
• Use X and Y scale bars as a nondestructive overlay
• Work with intensity or spatial measurements
• Create and display spatial calibration markers
• Create and display system calibrations
• Use pre-defined spatial calibration units
• Save and recall all calibrations
• Calculate derived calibrations automatically
• Store information about your microscope in lens information files

Metrology Tools
Metrology Tools Measure features such as best-fit line, arc, and circle.

Best-fit circle measurements

Measurements
Measure the lengths, areas, perimeters, and angles of objects in your image.

Angle measurements

Calculate Thickness
Determine the thickness of objects by calculating the mean, minimum and maximum thickness between lines.

Trace Objects
Use the trace tool to highlight dark features on a primarily light background, such as fingerprint ridges. These features can be drawn manually or automatically. The automatic tracing tool works by tracing the skeleton of dark features.

Automatic tracing tool

Detect Edges
Use the Caliper Tool to detect edges and calculate repeated measurements.

Caliper tool is used to count gear teeth.

Colocalization of Probes
The colocalization tools in Image-Pro Plus provide a visual representation as to the location and degree of overlap between two wavelengths.

Original fluorescence image	Area of interest (AOI) within the scatterplot selects the colocalized areas (orange & yellow)
Mask shows overlapping regions of red and green	Composite image showing FITC-labeled fibroblasts (green), Texas Red-labeled mitochondria (red)

Track Moving Objects
Manually or automatically follow cells or other organisms as they move through time and space. Use correlation tracking to follow objects when image segmentation is difficult or not possible.

Track and graph the changing of intensity parameters over time within an area of interest

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3D Rendering & Measurement

Explore the depth of your images by visualizing and interacting with them in three dimensions.

• View volumes from any angle, with adjustable transparency and shading.
• Zoom, rotate, and pan image stacks.
• Use adjustable lighting controls for enhanced viewing.
• Visualize clipping planes in orthogonal and oblique plane-selections.
• View internal features of 3D stacks by selecting volume of interest (VOI).
• Display synchronized, orthogonal views through an image stack with Stack Slicer.
• View multiple iso-surface renderings based on segmentation of selected intensities and colors.
• Project a virtual shadow of your image to get a better visual understanding of your object's 3D shape.

Apply Measurements to 3D Images
Obtain quantifiable data from your three-dimensional image through manual and volume measurements.

Industrial image created with Image-Pro 3D Suite - Images are courtesy of Ralph E. Napolitano of Ames Laboratory Solidification Science. Department of Materials Science and Engineering, Iowa State University

3D Manual Measurements:

• Individual point
• Point-to-Point distance
• Point-to-Surface distance line
• Line
• Angle
• Point-to-Line distance
• Circumference of an object bisected by plane
• Surface distance between two points

3D rendering of a kidney glomerulus - Image courtesy of Dr. Brian Matsumoto -Department of Molecular, Cellular and Developmental Biology University of California, Santa Barbara, CA

3D Volume Measurements:

• Volume
• Surface Area
• Sphericity
• Feret measurements
• Bounding box measurements
• Centroid information
• Automatic update of measurements from frame-to-frame playback of 4D sequences
• Set filter ranges for any measurements
• Clean borders

Track Objects in a 4D Sequence
Track and measure the movement of individual objects in a 4D sequence with the 4D tracking feature in 3D Constructor. Choose to automatically find 4D tracks in your image or manually track objects.

Visualize, Trace, and Count Neurons
Analyze neurons and other filament structures with the neuron tracing tool in 3D Constructor. Visualize and count the number of branches in a neuron as well as the length and volume of each branch.

Dendritic spine tracing of 3D neuron - Image courtesy of Ling Wang and Brendan Brinkman, University of California, San Diego, CA, USA

Explore Wavelength Overlap with 3D Colocalization
Use Image-Pro's colocalization tool to indicate which fluorescent regions are to be considered as overlapped, or colocalized. Then activate 3D Constructor to display the colocalized regions in three dimensions.

Create Animations of 3D Images
Share your 3D rendered images with others using 3D Constructor's easy 3D animation creation tools. Create animations with changing transparencies, slide positions, colors, etc. to give others a better understanding of the depth of your 3D image. Save in AVI or multi-frame TIFF or SEQ formats.

Display and Export Image Data
Display and interact with measurement data from individual, histogram, or scatterplot displays. Export 3D measurement data to Microsoft® Excel.

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Surface Analysis

Using only the in-focus information derived from a Z-stack of images, 3D Surface Inspector offers unique 3D views and measurements that reveal quality details. With a single camera and Z-motor, you will enjoy this cost-effective and illuminating analysis tool.

Using extended depth of field information, 3D Surface Inspector allows you to take inspection to the next level. Add 3D rendering and measurement to your toolkit and gain a much greater understanding of your specimens and samples.

Visualize Image Surfaces in Three Dimensions

Explore the depth of your images by visualizing and interacting with them in three dimensions.

• View the surface from any angle, using texture mapping, wire frame or Pseudo coloring to enhance details for interpretation
• Rotate the rendering actively to appreciate its structure
• Display the axes with spatially calibrated units
• Use adjustable lighting, contrast and shadow controls for enhanced viewing.
• Easily create AVI animations to share online or with others

Perform Manual Measurements:

• Differences in Z height between two X/Y positions
• Maximum and minimum Z values over an X/Y line profile
• Z height representation over an X/Y line profile
• Obtain surface area measurements for distinct object on a focal plane
• Measure the volume covered by that surface area

Surface area measurements of a coin

Choose Your Method of Image Acquisition

• Control a Prior Scientific™ Z-motor device directly
• Collect Z-stacks directly from Scope-Pro
• Import a series of Z interval images

Acquire Focal Planes Directly with Hardware Control

Control a Prior Scientific Z-motor stage directly from within the application

• The entire process is handled from within the application
• Alternatively, use Scope-Pro for Z-stack acquisition from other devices or stereo microscopes

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Automate, Customize & Share

Automate with Macros

Record - Save a series of Image-Pro commands as you perform them using macro recording tools.

Edit - Image-Pro Plus offers convenient customization tools and a built-in programming language to streamline your imaging research.

• Organize and implement macros in the Macro Browser
• Work with example and “Extras” macros included in Image-Pro Plus and Image-Pro Analyzer
• Use the IPBASIC engine with debugger, full editor, and dialog builder
• Record sessions and play them back in Basic Syntax
• Assign scripts to files and function keys
• Add basic loops and conditionals
• Edit and customize dialog boxes

For more advanced customization, Image-Pro Plus’ built-in IP Basic programming engine offers a debugger, full editor, and dialog builder. Use with COM objects such as Microsoft Word or Excel, and integrate and customize macros with Microsoft Visual Basic or Visual C++.

Share & Download Macros
The Media Cybernetics’ Solutions Zone is a searchable on-line database of macros, device drivers, plug-in modules, application programs and development consultants that support Media Cybernetics image analysis products. Visit the Solutions Zone to search for Image-Pro Plus macros in your industry.

Annotate
Point out features of interest and add text to your images with Annotation overlay tools.

Point out features of interest and add notes to your images with Annotation tools

Report and Publish Your Results
Export image data via DDE to Origin® and Microsoft Excel. Create custom reports with images data and text.

Create AVI Movies
Create AVI Movies Easily convert sequence files and 3D rendered images to AVI files.

Convert image sequences and 3D renderings to AVI files

Customize Your Workflow
Image-Pro Plus includes a variety of features that allow you alter the application to fit your workflow. Create a personalized Workflow Toolbar with buttons for your most frequently used tools.

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New Image-Pro Plus 7.0

Don't miss out on these excellent upgrade opportunities!

Image-Pro Plus Version 7.0 includes enhancements that speed up your acquisition and analysis tasks.

New Features in Image-Pro Plus 7.0 Include:

Microscope & Stage Automation Included

Image-Pro Plus includes microscope and stage automation tools which were previously available as add-ons. Control your microscope stage in any X, Y, or Z direction. Control motorized filter wheels, shutters, filter sliders, and lamp settings.

Accelerated Hardware Automation

Automation speeds have been accelerated for the following microscopy hardware controllers:

• Prior ProScan Controllers
• Olympus Microscope Stands (BX-61 and IX-81)
• Sutter Lambda 10-3
• Ludl MAC

Just-in-Time Image Loading

Save time by loading your multi-frame image sets on demand, resulting in extremely fast load times. This feature is available for Image-Pro sets and TIFF sequences.

Optional Live Tiling and Extended Depth of Field

Tile large images and perform Extended Depth of Field operations live – while acquiring your images. This tool is available as an add-on to Image-Pro Plus 7.0.

Live tiling image courtesy of Joop van Heerikhuize, Netherlands Institute for Neuroscience

Optional 3D Surface Analysis Tools

Use the optional 3D Surface Inspector tool to create beautifully illuminated 3D visualizations of your image stacks. Explore image depth with manual measurements and easily create .avi animations.

Use 3D Surface Inspector to analyze the surface area of objects

Visualize image depth with 3D Surface Inspectors' surface plot tool

Time-Saving Macros and a Macro Browser

A new Macro Browser (formerly the Macro Player) offers quick access to a variety of timesaving macros. New macros are also available in the Macros Menu.

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