I am currently researching image manipulation in the life sciences and stumbled upon a Western Blot looking very similar to the image below.

For creating the image I have cut out the blot using the Fuzzy Select Tool of GIMP and then greatly adjusted the local contrast of the blot as well as the background. By using the Fuzzy Select Tool some pixels of the original background are also captured. That is why the blot shown has a brighter background around the edges. I normally would not expect to see bright edges around a blot.

While I know how Western Blots look when they are captured with a digital camera, I do not know about techniques like Chemiluminescent Blotting (I am an undergraduate student in Computer Science). Maybe there is a blotting technique that results in an image like the one i created.

Is it possible to get a Western Blot result like this without inappropriate modification of the image?

imitated part Western Blot

  • $\begingroup$ Can you explain a bit more? I'm still confused by what that image is, in particular why it's colored the way it is. $\endgroup$
    – Amory
    Commented Oct 28, 2013 at 18:14
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    $\begingroup$ this looks like a normal band in a gel, with the image manipulated to replace every pixel above a certain cutoff to black. could be manipulation or it could be an accidental data issue $\endgroup$
    – shigeta
    Commented Oct 28, 2013 at 18:26
  • $\begingroup$ @Amory I have edited the question and provided some more details. Hope this helps. $\endgroup$ Commented Oct 28, 2013 at 18:39
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    $\begingroup$ Yeah I'm with @shigeta on this. Do you have the unaltered image? I can post an example of one of my chemiluminescent blots if you like. $\endgroup$
    – Amory
    Commented Oct 28, 2013 at 18:45
  • $\begingroup$ I have the unaltered image but i am not sure if it helps as the image posted is altered to imitate another image from an actual publication. The image i stumbled upon is from an actual publication and i can't post it here due to legal risks (but it looks almost exactly like the one i posted). I would appreciate to see an example of a chemiluminescent blot. $\endgroup$ Commented Oct 28, 2013 at 18:57

2 Answers 2


I do not think that a publication quality blot should have such an artifact, but I was able to find something similar by purposely over blotting (not the same as over exposing) a gel. If you use too much primary, secondary, or developing reagent, you can get your HRP signal to "burn in" a membrane where you get a distinct "negative band."

By negative band I mean a region that is more clear/white than background. This happens mostly when you have way too much secondary antibody. Normally when this happens, the CENTER of the band burns in first so you would see a white center with dark edges. And that's only when you are imaging the membrane as it is burning in. Often the whole band is white.

After going through the westerns of grad-student who is working with me, I found this image:

enter image description here

It came with the clear notation that it was burned in and disregarded (said student was learning how to do Westerns at the time). It's a GAPDH blot for those that care. When this does happen, it's obvious because you can then see the band with your naked eye on the membrane. Again this would be a reason to not publish the blot.

The image could have non-maliciously been edit via compression, screens, filters, etc as mentioned by others in the comments. This is the only way I can think of to experimentally have a similar image. Ether way I don't think the image should be reported as is.

  • $\begingroup$ Thank you, that's close to the image i posted. Still the white area is bigger and does not have sharp edges as the one posted. Assuming that one will not publish damaged blots the remaining option is that the image is edited inappropriately (with or without malicious intent). $\endgroup$ Commented Nov 4, 2013 at 20:10
  • $\begingroup$ @RalfThaenert as I mentioned it's similar, and I think as close as you can get to your image experimentally. I could imagine that if you, by immense chance, got an image just as the outer edge was fading you could get tighter, but it seems improbable. As scientists we are not prone to saying something is impossible, but rather that certain things are likely, especially compared to others. So yes, I think everyone thinks the simplest way to your image is digital editing (intended or not). $\endgroup$
    – Atl LED
    Commented Nov 5, 2013 at 1:27

Here's an example of a chemiluminescent blot:

enter image description here

For those who care, the first column on the left is the weight standard, the next two are pooled pellets, and the rest are increasing fractions of purified HIV using a mouse monoclonal anti-p24 (capsid) antibody.

This blot is pretty clean (although for my work it was pretty disappointing...) but here's one that looks less-than-ideal, and perhaps more like your example:

enter image description here

These are lysates and pelleted supernatant from Jurkat uninfected (left) and HIVIIIB-infected (right) cultures, detected with a mouse monoclonal anti-tsg101 antibody.

I agree with shigeta's comment that your image might not be exactly what I would expect, but that doesn't necessarily mean it was done maliciously; as you can see, there can be a fair amount of variation in how blots turn out depending on the equipment used, the type of membrane, the protein itself, and so on. I try to be pretty conservative with image filters but there's a lot you can do with those guys that can be shady but falls short of Photoshop.

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    $\begingroup$ Thanks for the images. The images do not show such a behaviour around the blot regions. So there is no Western Blot technique which shows this behaviour by default? Of course such an image can be the result of compression, filtering and beautification and is not necessarily an intended manipulation. $\endgroup$ Commented Oct 28, 2013 at 19:57

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