In the end, the integrity and robustness of the published scientific data should be part of the considerations while planning image analysis experiments. Sometimes, a qualitative interpretation might be better suitable than a flawed quantification. IMHO: Not because we can measure everything means also we should do it. For the complete list of video tutorials and support documents, visit Image Studio Support. These include such functions as quantification, molecular weight determination, and normalization. I can only recommend to use either the software of the Western Blot imaging system in the lab or alternatively GelAnalyzer because it allows to stick pretty well to the recommended procedure by Western Blot material suppliers (which I would guess are the most experienced people in that particular field) and as seen in the publication above. This video tutorial demonstrates the Image Studio 5.x Western Analysis Ribbon, which provides tools to analyze Western blots. Hammond, “A defined methodology for reliable quantification of Western blot data.,” Mol. This protocol offers an Image J macro/plugin that enable easy quantification of bands on western blots, dot blots, and fluorescent gels etc. So, generally there are many pitfalls related to WB measurements.īesides the literature list in one of the linked posts above, mainly the following gives a good insight into the procedure: The same technique can be used for quantification of DNA or RNA from films. And band selections which overshoot the actual band also lead to wrong results. One cannot reliably quantify bands by making boxes around them and using analyze measure nor do horizontal lines fulfill the requirements. Here is one video which is explaining a little more detailed the considerations of WB in general, normalization as well as measurements Western blotting is routinely used to detect proteins and their posttranslational modifications (PTM) in biological samples. Normalization factors must be calculated for each blot do not use the same normalization factors among different blots. The latter and pretty much of most other ones completely ignore every thing mentioned in scientific literature regarding Western Blot measurements (and the pre-requisites for it). Data analysisnormalizing your western blot data Once a housekeeping protein has been validated, it can be used to normalize the data. ![]() There are many videos online like the linked one above. ![]() But I cannot contain myself to add my 2 cents to this topic, because some of those videos make me like… No offense to no one making those videos or taking them as orientation in case of the lack of other available resources. ![]() This was the one I looked at Analysing blots and gels with ImageJ/Fiji - YouTube
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |