My lab has recently found that even when our ponceau stains look homogeneous our single protein loafing controls such as GAPDH or pan actin have been highly variable after harsh stripping. ¹This subreddit is not affiliated with the creators of ImageJ or FIJI, but is simply a place to share ideas, papers, resources, and expertise - especially as relate to questions & answers posted here. Sign-up for one of the mailing lists: /Mailing_Lists It also hosts a forum for interacting with the developers.įIJI Is Just ImageJ - "a distribution of ImageJ (and ImageJ2) together with Java, Java3D and a lot of plugins organized into a coherent menu structure." is full of ImageJ development and analysis resources. Image analysis is interdisciplinary, so clearly explain field-specific terms or jargon. Clearly explain what you are trying to learn, not just the method used, to avoid the XY problem. Provide details: Be thorough in outlining the question(s) that you are trying to answer.People from the future may be stuck trying to answer the same question. Report spam or content that is hateful or off-topic.Upvote those who contribute to the discussion and provide freely of their time to assist you.Projects: Share a Link to your pet image analysis project.Research: Links to published (articles in scientific journals or in established repositories) that utilize ImageJ/FIJI for image analysis or are about image analysis.Discussions: Text posts, meant to ask about general issues relating to image analysis.Image analyst job posts are also welcome. Tips: Text or Link posts to share useful how-to tricks and discoveries on using ImageJ/FIJI.Questions which have been Solved will be marked as such. This could include algorithms, microscopy and scientific imaging, plug-ins, methods, and specific features of the software. Questions: Text posts asking about image analysis and ImageJ/FIJI.This is a an unofficial¹ forum to discuss image analysis, software features, to get help, to share ideas, and to share work done using ImageJ or FIJI. It's used worldwide, by a broad range of scientists. It also produces publishable figures which indicate exactly the quantification areas which is extremely important for a transparent measurement.ImageJ is a freely available, open source image processing and analysis program using Java, on which FIJI is based. I will try to get in contact with the authors and see if there might be a possibility to link it as plugin into ImageJ (e.g. It is freely available but not completely open source with access to the source code. The design of a quantitative western blot experimentģ.) Since I first tried to code a plugin which would combine the necessary functions for ImageJ, I stumbled over an existing and extremely good tool (also Java) with all the things you would need to run such an analysis (including calibration).Western Blotting Inaccuracies with Unverified Antibodies: Need for a Western Blotting Minimal Reporting Standard (WBMRS).Evaluating Strategies to Normalise Biological Replicates of Western Blot Data.A Defined Methodology for Reliable Quantification of Western Blot Data (Here you see the analysis methos indicated).Quantifying Western blots: Pitfalls of densitometry.So, here a few recommendations and very helpful links (since this is not my invention):ġ.) have a look at the following video seminar from Aldrin Gomes as an introduction to the problem on the experimental side:Ģ.) Here a few publications which help in getting a better understanding before the actial image analysis So, even while often suggested, drawing boxes around a band is not the proper way! 2: The ►Analyze ►Gel tools and the ►Analyze ►Calibrate… function provide theoretically everything what you would need but a clear guideline is still missing. That’s why I posted it here!Įverything I have found in the web so far was mostly going a in the wrong direction leading to non-reliable measurements! 1: Many students try to find a guideline for Western blot “quantification” online while using ImageJ. And I think the topic is important in the aspect of reliability and reproducibility in science. So the following topic is not directly linked to ImageJ necessarily but many of this community might still be interested in. Since once in a while the topic semi-quantitative Western blot analysis pops up and many people in biological sciences who do Western blots or gels are into that, I wanted to share some information on that.
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