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The dangers of overexposing western blots

It’s like, how much more black could this be? And the answer is none. None more black.

Nigel Tufnel, lead guitarist of Spinal Tap.


None more black (or none more white) is a real concern when you’re talking about imaging western blots. If you only want to verify the presence of a protein, you can stop reading here, because even an overexposed image will tell you that there is some protein present in your sample. But an overexposed image of a western blot is essentially useless when you’re … Continue Reading

ImageJ StartupMacros

Before I lose this file again: StartupMacros.txt This is a text file, based on the original StartupMacros.txt file that came with ImageJ, that can be dropped in the ImageJ/macros folder that is created when you install ImageJ. Start up ImageJ, and if this macro doesn’t automatically run, go to Plugins>Macros>Startup Macros to load it. It installs a few more drawing tools on the ImageJ toolbar and creates some keyboard shortcuts for the following tools:

  • Line tool – press ‘l’
  • Brush tool – press ‘b’
  • Eraser tool – press ‘e’
  • Wand tool – press ‘w’

I either found or created some code … Continue Reading

Digitizing data from old figures with ImageJ

Following on this earlier post concerning the retrieval of data from figures in published papers with R, here is a method for doing the same with ImageJ. This is a freely available program that should run on Windows/Mac/Linux, just like R. To extract data points from a figure, you’ll need the Figure_Calibration plugin developed by Frederic V. Hessman, available here. Download the Figure_Calibration.class file from that page and save it to your ImageJ/plugins/ directory. Then load up ImageJ.

[caption id=”attachment_1051″ align=”aligncenter” width=”400″ caption=”The example figure that we wish to retrieve data from. We can use … Continue Reading

ImageJ paint brush tool keyboard shortcut

ImageJ is easy to modify if you know a bit of Java. I don’t know any Java. But here’s what I did to add keyboard shortcuts for the paint brush tool and the line selection tool. The code is based off this help file from the ImageJ website: http://rsbweb.nih.gov/ij/docs/guide/userguide-32.html

The first step was to figure out to refer to the line tool and paint brush tools. I did this by going to Plugins>Macros>Record and clicking on the line tool and the paint brush tool in succession. The numeric codes for those tools were revealed in the macro recording window … Continue Reading

Measuring leaf or algae blade area in a digital photo with ImageJ

The following is a tutorial on using the freely available ImageJ program from the National Institutes of Health for estimating the area of a leaf or algae blade (or anything else). The process is straightforward once you understand the various steps.

You must begin with a photograph of your object, flattened and dried off as much as practical. Include a clearly visible ruler or other measuring device in the picture. Ideally you will photograph your dark leaf or blade on a light background to maximize the contrast between the object and the background.

[caption id=”attachment_1009″ align=”aligncenter” width=”300″ caption=”The original picture, … Continue Reading

Analyzing gels and western blots with ImageJ

The following information is an updated version of a method for using ImageJ to analyze western blots from a now-deprecated older page.

If you’re looking for a more comprehensive workflow option for your western blot analyses, please visit my tutorial on using Image Studio LiteIS_lite_icon, a free software package from LI-COR Biosciences. The Image Studio Lite software can be downloaded for free from www.licor.com/islite. In addition to the functionality described below for ImageJ, Image Studio Lite provides additional … Continue Reading