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The simplest method to convert to grayscale is to go to Image>Type>8-bit. The gel analysis routine requires the image to be a gray-scale image. This version of the tutorial was created using ImageJ 1.42q on a Windows 7 64-bit install.Ģ. There should be very little difference between the results obtained from the various methods. You may prefer to use it instead of the methods I outline below. The method outlined here uses the Gel Analysis method outlined in the ImageJ documentation: Gel Analysis. For an explanation of why overexposure will ruin your results, see this page. This also assumes that you didn’t overexposed your blot image when you produced it. See the references at the end of this tutorial for a discussion of the various ways that you can screw this step up.

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film), and use the scanner software to save 16-bit tiff images. If you are scanning x-ray film on a flatbed scanner, make sure you use a scanner with the ability to scan transparencies (i.e. tif would be the preferred format to retain the maximum amount of information in the original image). This tutorial assumes that you have carried your gel or blot through the visualization step, so that you have a digital image of your gel in. ImageJ ( ) can be used to compare the density (aka intensity) of bands on an agar gel or western blot.

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The Image Studio Lite software can be downloaded for free from In addition to the functionality described below for ImageJ, Image Studio Lite provides additional data management features along with annotation tools for producing publication-ready images of your westerns. If you’re looking for a more comprehensive workflow option for your western blot analyses, please visit my tutorial on using Image Studio Lite, a free software package from LI-COR Biosciences. Plant Proteases and Plant Cell Death: Methods and Protocols. Detection of Damage-Activated Metacaspase Activity by Western Blot in Plants. If you need a peer-reviewed citation for the methods outlined below, you may cite Stael, S., L.P. The following information is an updated version of a method for using ImageJ to analyze western blots from a now-deprecated older page.

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