Since the advent of digital imaging, inappropriate manipulation of images has become a significant problem in scientific publishing.  Authors need to understand what is an acceptable enhancement for purposes of clarity or emphasis versus publishing misconduct.  Undocumented image manipulations can lead to accusations of research misconduct.  The Office of Research Integrity (ORI) from the U.S. Department of Health & Human Services stated that 67% of their closed research misconduct cases between 2011 and 2015 involved image manipulation.

Other ORI guidance includes:

• Clearly document all changes made to an image.
• Retain the unprocessed image for your records.
• Follow journal guidelines for permissible processing.

The Rockefeller University Press has established 4 basic guidelines:

• No specific feature within an image may be enhanced, obscured, moved, removed, or introduced.
• Adjustments of brightness, contrast, or color balance are acceptable if they are applied to the whole image and as long as they do not obscure, eliminate, or misrepresent any information present in the original.
• The grouping of images from different parts of the same gel, or from different gels, fields, or exposures must be made explicit by the arrangement of the figure (e.g., dividing lines) and in the text of the figure legend.
• If the original data cannot be produced by an author when asked to provide it, acceptance of the manuscript may be revoked.

Editorial guidelines from the Proceedings of the National Academy of Sciences of the United States of America (PNAS):

No specific feature within an image may be enhanced, obscured, moved, removed, or introduced. The grouping or consolidation of images from multiple sources must be made explicit by the arrangement of the figure and in the figure legend. Adjustments of brightness, contrast, or color balance are acceptable if they are applied to the whole image and if they do not obscure, eliminate, or misrepresent any information present in the original, including backgrounds.

Questions about images raised during image screening will be referred to the editors, who may request the original data from the authors for comparison with the prepared figures. If the original data cannot be produced, the manuscript may be rejected. Cases of deliberate misrepresentation of data will result in rejection of the paper and will be reported to the corresponding author's home institution or funding agency. Authors must obtain consent for publication of figures with recognizable human faces.

• Cromey D.W. (2012) Digital Images Are Data: And Should Be Treated as Such. In: Taatjes D., Roth J. (eds) Cell Imaging Techniques. Methods in Molecular Biology (Methods and Protocols), vol 931. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-056-4_1 Detailed information about all aspects of capturing and processing images for both analysis and illustration.  It is written for biologists but the extensive background is useful for a range of scientific disciplines.  

• Microscopy Society of America (MSA) - Policy on Digital Imaging https://www.microscopy.org/resources/digital_imaging.cfm  

• Also see Science Integrity Digest by Elisabeth Bik or follow her on Twitter for real time image manipulation case studies. https://scienceintegritydigest.com/how-to-guides/ 

• Jambor H, Antonietti A, Alicea B, Audisio TL, Auer S, et al. (2021) Creating clear and informative image-based figures for scientific publications. PLOS Biology 19(3): e3001161. https://doi.org/10.1371/journal.pbio.3001161  Biology focused but has good suggestions for generating images for publication.  

• Data Curation Network Primer on Confocal Microscopy - tutorial on management of microscopy images and metadata.

• https://www.councilscienceeditors.org/resource-library/editorial-policies/white-paper-on-publication-ethics/3-4-digital-images-and-misconduct/ 

• https://www.pnas.org/author-center/editorial-and-journal-policies#image-integrity - Guidelines for image integrity from the Proceedings of the National Academy of Sciences.