8/15/2023 0 Comments Imagej particle analysis![]() With the line still highlighted by the threshold widget, run Analyze ▷Tools ▷ Analyze Line Graph↑ to get the XY coordinates of the traced line (you can hold down Alt while selecting the command to reveal the actual image that is processed ).Select the curve with the Wand Tool↑ and run Edit ▷ Clear Outside↑ to erase everything on the canvas but the curve.Open the Threshold… ↑ tool ( Shift T) and adjust the threshold levels so that the curve is highlighted in red.Alternatively, use one of the drawing tools ( Pencil↑ or Brush↑) to paint directly in background color. Use any of the Area Selection Tools↑ as an eraser (press Backspace to erase) in order to isolate the single graph curve to be measured. Set background color to white using the Color Picker Tool↑ or the Color Picker window↑.Analyze Line Graph will assume that the graph is displayed on a white background so images with darker backgrounds must be adjusted beforehand ( see e.g., Edit ▷ Invert ↑ and Process ▷Binary ▷ Make Binary↑). Make sure your graph is a grayscale image ( Image ▷ Type ▷↑8-bit). For practice, use the File ▷ Open Samples ▷↑Line Graph (21K) sample image. The following procedure describes how to use it: This command uses the Particle Analyzer↑ to extract sets of coordinate data from digitized line graphs. Select Gels ▷↑Label Peaks to label each measured peak with its size as a percent of the total size of the measured peaks.If necessary, scroll the image vertically by holding down the space bar and dragging. For each peak, measure the size by clicking inside the peak with the Wand Tool↑.To access to all the lanes, it may be necessary to scroll the image vertically using the Scrolling Tool↑ (Hold down the space bar to temporarily switch to this tool). Note that you can hold Shift to constrain lines to be either horizontal or vertical. Use the Straight Line Selection Tool↑ to draw base lines and/or drop lines so that each peak of interest defines a closed area (ImageJ will automatically switch to the Straight Line tool).Select Gels ▷↑Plot Lanes ( 3 ) to generate the lane profile plots.Repeat the previous step for each remaining lane.The selected lane is outlined and labelled, and ‘Lane n selected’ is displayed in the status bar. Move the rectangular selection right to the next lane (or down if the lanes are horizontal) and select Gels ▷↑Select Next Lane ( 2 ).Select Gels ▷↑Select First Lane ( 1 ) and the lane will be outlined and ‘Lane 1 selected’ displayed in the status bar.This should be the left most lane if the lanes are vertical or the top lane if the lanes are horizontal. Use the rectangular selection tool to outline the first lane.Note that a copy of the gel image with the lane outlines can be created at any point using the Image ▷Overlay ▷ Flatten ↑ command. Responsible for Icy's kernel architecture, software code and website.For practice, refer to the video tutorial on the ImageJ wikipage and use the File ▷ Open Samples ▷↑Gel sample image (1-D gel) to perform the following steps. Researcher at the quantitative analysis unity at the Pasteur Institute, Paris. Working with ImageJ/FIJI for microscopy image analysis and teaching. Head Software developer at Biomedical Imaging Group (BIG) at Ecole Polytechnique Fédérale de Lausanne (EPFL). A tutorial on ImageJ/FIJI plugin programming will also take place together with scripting and graphical programming for ICY. In this context, we will focus on the basic operation of both ImageJ/FIJI and ICY as well as several plugins for the solution of common issues in image analysis such as segmentation, particle analysis, co-localization, deconvolution, spot count and particle tracking. ICY is an open source software that is compatible with ImageJ plugins but which enables other functionalities. ImageJ/FIJI are based on an open source architecture, which enables functionality extensibility using plugins and Macros. ImageJ, its distribution Fiji and ICY are widely-used public-domain software for microscopy image analysis. This hands-on workshop is directed to users working on biology and biomedicine that require objective analysis of microscopy image data.
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