Blind spots on western blots: assessment of common problems in western blot figures with recommendations to improve them

This project contains protocols, data and code for a meta-research project examining the prevalence of image display, data presentation and methodological reporting practices in original research articles in cell biology and neurosciences that include western blots. Papers were identified from the top 25% of journals in each field

Number of articles examined by journal (cell biology).

Values are n, or n (% of all articles). Articles that were not full-length original research articles (reviews, editorials, perspectives, commentaries, letters to the editor, short communications, etc.) or did not include eligible images were excluded. (DOCX)</p

Western blot: From gel to publication.

Western blotting is a standard laboratory method that uses antibodies to detect target proteins in a sample. (1) The sample, typically a mixture of proteins, is loaded on the gel. A molecular weight (MW) marker, which contains prelabeled proteins of varied, known molecular weights, is loaded on the gel alongside the protein sample as a size reference. (2) Gel electrophoresis is used to separate proteins based on their molecular weight. (3) The proteins are transferred, or “blotted”, onto a membrane. (4) The membrane is blocked to reduce nonspecific binding and then sequentially probed with a primary antibody that specifically binds to the protein of interest and a secondary antibody. The latter binds the primary antibody and carries an enzyme or a fluorophore that allows subsequent detection. (5) The signal is detected through a chemiluminescent reaction or fluorescence, respectively. (6) An image of the western blot is prepared for publication: Annotations are added and often the blot is cropped. For the unprocessed image, see S1 Fig.</p

Western blot image minimal reporting standard.

Published western blots should show a minimum of 2 molecular weight marker bands of different weights if the protein of interest falls between the molecular weight marker bands, and 3 molecular weight marker bands of different weights if the protein of interest is directly at one of the marker bands. The molecular weight markers should be annotated with labels. An original, uncropped image of each blot should be published in the supplement or deposited on a public repository. The source data blot should be named in a way that links it to a specific figure, panel, and protein. The outline of the crop should be annotated on the original image. For the unprocessed image, see S1 Fig.</p