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Gel Diffusion Image Analysis Neil Forbes Nanotechnology Institute 2007.

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Presentation on theme: "Gel Diffusion Image Analysis Neil Forbes Nanotechnology Institute 2007."— Presentation transcript:

1 Gel Diffusion Image Analysis Neil Forbes Nanotechnology Institute 2007

2 Software Download ImageJ –Install files is available at: –Follow the Downloads link –Downloads are available for Windows, Macintosh and Linux Download Radial profiler –http://rsb.info.nih.gov/ij/plugins/radial-profile.htmlhttp://rsb.info.nih.gov/ij/plugins/radial-profile.html –Or in ImageJ: Help ImageJ Website Plugins Radial Profile Plot Radial Profile Plot –Download the plugin to the plugins folder in Image Program File –Restart ImageJ

3 Image Analysis Three methods from simplest to more complicated Method 1: –Measure the dye penetration distance each day using a ruler –Use graph paper to plot distance vs. time –The rate is the slope of the line

4 Image Analysis Method 2: –Display all of the images on the computer –Distances in the images need to be calibrated –Use the line function to measure the dye penetration distance (D) in pixels –Use a ruler to measure the width of the gel (W) in pixels, which we know is 60mm –The penetration distance is: –Plot the distance vs. time

5 Measurement by Hand

6 Image Analysis Method 3: –Open all of the images –Split into (R)ed, (G)reen and (B)lue images (RGB Split) –Pick the image with the most information –Draw a square box around the gel –Apply Radial profile from the plugins menu –Save the profile

7 Image Analysis Method 3 (cont): –Open all of the profiles in a spreadsheet (Excel) –Normalize the intensity: –Find the pixel that crosses 0.5 –Divide by the total number of pixels and multiply by 30 mm (the gel radius) –Plot distance vs. time

8 Software Analysis

9 Comparison – Pretty good! The dye penetrates at approximately 200 μm/hr

10 Image Analysis Finally, if the distance is normalized, complete distance vs. time plots can be generated. These compare nicely to theoretical predictions.


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