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Angiogenesis Analyzer for ImageJ

Saturday 20 October 2012, by Gilles Carpentier

  Table of contents  


The different functions are accessible from the tool bar of ImageJ. They can be grouped in menu tools to simplify the use of this program.

  • "Network Analysis Menu Tool":

    • "Analyze HUVEC Phase Contrast" and "Analyze HUVEC Fluo" functions, allow the network analysis organization of individual HUVEC images taken in phase contrast or in fluorescence. Note that to avoid confusion, phase contrast images have to be encoded in RGB color 24 bits and fluorescence images in 8 or 16 bits.
      Below: summary for the results of these functions - slide-show:
      HUVEC Phase Contrast HUVEC Phase Contrast + Analysis HUVEC Fluo HUVEC Fluo + Analysis
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    • "Analyze Binary Tree" performs the analysis of the network organization of a skeleton (or tree) in a binary image. This function is useful for the tuning of a new set of experiments. Further analysis must be performed with "natural" cell images.
      Below, binary tree analysis - slide-show:
      Binary tree Analyzed tree
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    • "Save Current Analysis", "Close Current Analysis" and "Save Current Analysis and Close" Save (and optionally close) at the initial image file location the current analysis (images and results table) in a folder with the name of the image with the suffix "=AN".
    • "Blink Overlay (b)", "Hide Overlay (h)" and "Show Overlay (s)" Use these functions to evaluate the pertinence of the results by making blink the overlays corresponding to the detected network elements. Use the keyboard shortcuts "h" and "s" to control manually the blinking.
  • "Blurred Mask Tool":

    This tool removes the gradient in a selection of a phase contrast image. It is convenient to remove some dirt which generates some artifactual network elements.

  • "Tuning Functions Menu Tool":

    This menu tool regroups several functions and settings to tune the analysis parameters.

    • "Find a Tree" return a binary skeleton (or tree) of a natural HUVEC image acquired in phase contrast or fluorescence.
    • "Find & Remove Loops" removes the artifactual loops in a binary tree image on the criteria of the size.
    • "Find Extremities" detects the extremities in a binary tree. It returns results as overlays and binary map.
    • "Find Nodes and Junctions" detects nodes (pixels with 3 neighbors) as a circular dot and junctions, which correspond to nodes or group of fusing nodes.
    • "Find Nodes and Branches" Performs the same of the two precedent functions (detects extremities, nodes and junctions) plus other elements of the ramification:
      • segments; elements delimited by two junctions.
      • branches; elements delimited by a junction and one extremity.
      • twigs; a twig is a branch whose size is lower than an user defined threshold value.
      • isolated elements are binary lines which are not branched.
    • "Record the Steps of Limbing" record in a dedicated repertory, the results of every iterations required to get an analysis corresponding to a limbing, including master segments, master junctions and meshes detection.
      • Master segments consist in peaces of tree delimited by two junctions none exclusively implicated with one branch, called master junctions.
      • Master junctions are junctions linking at least three master segments. Optionally, two close master junctions can be fused into an unique master junction.
      • Meshes are areas enclosed by segments or master segments.
    • Detection of constitutive elements of the network - summary. Below: extremities (arrow head A-B); nodes, identified as pixels that had at least 3 neighbour, corresponding to a bifurcation (arrow A-B); twig (C1, D1), segment (C2, D2) delimited by two junctions (C3, D3) (note that this pointed junction is composed by several nodes) and branch (C4, D4). E shows a junction implicated only in branch (E1) and master junctions like E2 delimiting master segments (E3). F shows the master tree composed from master segments associated by master junctions delimiting the meshes (F1). Optionally, two close master junctions can be fused into an unique master junction (F2). Note the underlying segment (F3).
      PNG - 351.2 kb
      Detection of constitutive elements of the network.

    • "Get Maps of Selections" Once analyzed an image is presented with different detected elements as overlayed selections. This function allows to build new maps of overlays with only user selected elements.
  • "Batch Image Treatment Tool":

    By selecting a folder with this tool, the program will analyze batch of images contained in multi level repertories. Results and time statistics are available during the treatment (see picture below).

  • "Measurement Table Manager Menu Tool":

    Menu tool allowing to open, modify and save "Stat Results Table".

  • "On Line Documentation & Demo Menu":

    On line ressources: documentation, phase contrast and fluorescence demo images for training.

  • "Version and Update Infos":

    Click on this ImageJ tool bar icon to look for new versions.

  • "About Angiogenesis Analyzer":

    Gives an off line short documentation.

logoij ImageJ ( is a public domain Java image processing program inspired by NIH Image for the Macintosh. It runs, either as an online applet or as a downloadable application, on any computer with a Java 1.1 or later virtual machine. Downloadable distributions are available for Windows, Mac OS, Mac OS X and Linux. The author, Wayne Rasband (, is at the Research Services Branch, National Institute of Mental Health, Bethesda, Maryland, USA.
Gilles Carpentier, Faculte des Sciences et Technologie,
Universite Paris Est Creteil Val-de-Marne, France.

Special thanks to Alessandra Albano for the English correction of these sites.
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