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Scientific images may be adjusted for clarity, but the edit must preserve the evidence. In practice, that means whole-image brightness, contrast, or color changes can be acceptable when they do not hide or invent information, comparable images and controls receive equivalent treatment, composites are clearly identified, processing is disclosed, and original files are retained. Local touch-ups, selective enhancement, erased or added features, and undisclosed composites cross the line. The target journal’s current instructions determine the submission details.
Start with the evidence-preservation rule
An image in a paper is data, not artwork. Processing is permissible when it improves legibility without changing what a reader could conclude from the underlying acquisition. That principle applies to microscope images, fluorescence channels, gels, western blots, radiographs, scans and other scientific figures.
There is no single universal rulebook. Nature Portfolio, Elsevier, IOP Publishing, Wiley guidance, KU Leuven and the Council of Science Editors (CSE) express a broadly shared standard, but differ on where disclosures belong, whether raw files must accompany a submission and how particular image types are handled. Treat the principles below as a practical baseline, then follow the live instructions for the journal you are submitting to.
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The following operations can be legitimate when applied in a way that keeps the original information interpretable.
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Whole-image brightness, contrast and color balance
Adjusting brightness, contrast or color balance across the entire image is commonly allowed if no relevant signal is concealed, removed or misrepresented. A global adjustment should not turn a weak band into an apparent strong band, make a background feature disappear or clip values that a reader needs to assess.
Apply equivalent processing to experimental images, controls and other images being compared. A global adjustment made only to an experimental panel can mislead even though the individual panel was not locally edited.
Limited cropping
Cropping can remove empty margins and focus a figure on the relevant region. The crop must not conceal context, remove a control or band needed for interpretation, or imply that the displayed region is the entire acquisition when it is not. For gels and blots, retain the bands and controls that support the stated conclusion and keep the uncropped scan.
Labels, arrows and scale information
Annotations such as panel labels, arrows and scale bars may be added to explain what the reader is seeing, provided they are clearly annotations and do not alter the underlying data. An arrow should point to a feature; it must not cover or replace one. A scale bar must reflect the acquisition and display scale rather than being decorative.
Pseudocolor and channel display changes
Pseudocolor can make intensity or channel information easier to read, especially in microscopy. Channel-specific color choices, lookup tables and merged-channel displays are acceptable in appropriate contexts when the figure and legend explain what colors represent. A rainbow lookup table, for example, can create visual boundaries that are not present in the data, so disclose the lookup table and any transformation that affects interpretation.
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Gamma, nonlinear transforms, deconvolution and projections
Nonlinear adjustments such as gamma changes, deconvolution, projections and related operations can be scientifically useful. They are not automatically prohibited, but they change how values are displayed and therefore require disclosure. Explain the operation, software and relevant settings, and do not present a processed result as though it were an untouched acquisition. If processing increases apparent resolution, state the measured resolution and describe downstream processing or averaging that contributed to the appearance.
| Operation | When it can be acceptable | What to record or disclose |
|---|---|---|
| Brightness, contrast or color balance | Applied globally, with information preserved and equivalent treatment of comparable images | Software, settings or method, and the fact that the adjustment was global |
| Crop | Removes irrelevant area without hiding context, controls or relevant signal | What was cropped when the crop affects interpretation; retain the full original |
| Arrows, labels and scale bars | Clearly separated annotations that do not modify data | Identify annotations in the figure or legend when needed for clarity |
| Pseudocolor or lookup table | Used to display channels or intensity in a scientifically meaningful way | Color mapping, lookup table and channel assignment |
| Gamma or other nonlinear transform | Used for a stated analytical or display purpose without changing the conclusion | Transformation, software and key parameters |
| Deconvolution, projection or averaging | Method is appropriate to the acquisition and interpretation | Method, software, settings, number of images or planes where relevant, and effects on apparent resolution |
Edits that compromise the evidence
The central prohibition is selective alteration of a specific feature. Nature Portfolio describes data beautification as unacceptable, and Elsevier states that a specific feature must not be enhanced, obscured, moved, removed or introduced. The same boundary appears in IOP and KU Leuven guidance.
- Cloning, healing or content-aware removal: Do not erase a blemish, cell, band, spot or background feature, and do not copy a feature into another location.
- Local enhancement: Do not brighten one cell, band or region to make it appear stronger than it was.
- Selective suppression: Do not darken or blur a region, remove noise locally or hide a background signal when that changes interpretation.
- Moving or rearranging evidence: Do not shift a feature or move a lane so that its position supports a different conclusion.
- Adding evidence: Do not introduce a band, cell, edge or signal that was not acquired.
- Extreme global adjustment: A whole-image operation is still unacceptable if clipping or thresholding makes relevant data disappear.
- Unequal treatment: Do not process an experimental image more strongly than its control or comparison image.
- Hidden composition: Do not join separate fields, gels, exposures or time points and present them as one continuous original image.
These restrictions apply even when the alteration is made for appearance rather than analysis. An edit that makes a figure look cleaner can still make the viewer reach a different conclusion.
Consistency is part of validity
Readers compare panels, not just individual images. Process images of the same type with equivalent steps and apply the same display logic to controls and experimental conditions. If one panel receives a stronger contrast stretch, a different gamma value or a different background treatment, the visual comparison is no longer fair.
Document the processing path so that another researcher can understand whether the images were handled in parallel. “Adjusted for clarity” is too vague when different images received different operations. State the software, the operation and the relevant settings, especially for nonlinear transforms, channel-specific adjustments and intensity projections.
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How to assemble a composite figure without misleading readers
A composite figure can combine panels from separate acquisitions, but the assembly must be visible and explained. Use dividing lines, spacing or another unmistakable boundary when parts come from different gel regions, gels, fields, exposures, locations or times. The legend should identify the source of each part and explain why the assembly was made.
Never imply that nonadjacent lanes were next to one another or that images from separate locations formed one continuous field. Nature cautions against combining images from different times or locations as a single image except when the result is explicitly described as a time-averaged or time-lapse representation.
Whether a particular composite is allowed depends on the experiment and journal. A visible boundary and a caption disclosure do not make an otherwise invalid comparison valid; they make the provenance of the displayed parts clear so an editor and reader can evaluate it.
What a composite legend should answer
- Which panels, lanes or regions came from which acquisition?
- Were the samples from the same experiment, parallel experiments or different runs?
- Were the source images processed in parallel?
- Were any lanes rearranged, and are the boundaries visible?
- Was a projection, averaging step, time combination or channel merge used?
Disclosure: what to record and where to say it
Keep a processing record while the figure is being made. Name the acquisition equipment and software, note important acquisition settings, and list each processing operation that affects display or interpretation. For microscopy, include channel-specific adjustments, pseudocolor or lookup-table information, gamma or other nonlinear changes, deconvolution, projection, averaging and measured resolution when those details matter.
Put the relevant description in the Methods. Add it to the figure caption or legend when the processing changes how the image should be read or when the journal requires caption-level disclosure. Some venues also ask for a declaration during submission. The exact location is journal-specific, so check the current author instructions rather than assuming that a Methods sentence is sufficient.
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Preserve the acquired files, metadata and a copy of the final figure. A flattened presentation file is not a substitute for the original. Editors may request the originals during peer review or a later assessment.
A practical pre-submission workflow
- Preserve the source data. Save unprocessed image files, acquisition metadata and the final figure in separate, traceable locations. Keep the original gel or blot scans and the full microscope fields, not only cropped exports.
- Define the comparison. Identify which images, controls and panels will be compared. Decide what equivalent processing means for that set before adjusting one panel.
- Use the least manipulative edit. Prefer a whole-image adjustment that improves legibility while retaining all relevant information. Avoid local tools and aggressive clipping.
- Check every feature against the source. Compare bands, cells, signals, edges and background regions with the original. Confirm that nothing relevant vanished and nothing was added or moved.
- Make assembly visible. Add dividing lines or another clear boundary for parts from different fields, gels, exposures or times. Do not let layout imply continuity that did not exist.
- Write the processing record. Record software, acquisition equipment, key settings, transformations, channel changes, projections, averaging and any rearranged lanes.
- Place disclosures correctly. Put the required details in Methods, the legend or caption and the submission declaration specified by the journal.
- Read the live journal instructions. Confirm rules for raw-image submission, microscopy details, gel and blot presentation, and the location and wording of disclosures before uploading the manuscript.
Microscopy: details that commonly require extra care
Nature Portfolio’s microscopy guidance illustrates why a simple “global versus local” rule is not enough. It advises applying adjustments to the entire image and avoiding threshold manipulation, expansion or contraction of signal ranges, and changes that alter high-intensity signals. If gamma, pseudocolor or individual channel adjustments are used, describe them.
For merged images, identify channel-specific changes so a reader can interpret displayed colors and relative intensities. State the processing software and, where relevant, the lookup table. Report measured resolution and downstream processing or averaging that enhances apparent resolution; otherwise a processed image can look more resolved than the acquisition supports.
A scale bar and arrows can aid interpretation, but they do not replace acquisition and calibration details. Keep the unprocessed field so the displayed crop and annotations can be checked.
Gels and western blots
Quantitative comparisons across separate gels or blots are especially difficult to interpret. Nature strongly discourages them. If a comparison across separate runs is unavoidable, the legend should state whether the samples came from the same experiment or parallel experiments and that processing was parallel.
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Rearranged nonadjacent lanes must have visible boundaries and an explicit legend explanation. A crop should retain important bands and controls, and the original scan should remain available. Do not use contrast or background treatment to make a faint band appear present or to make an unwanted band disappear.
Channel merges, projections and other display composites
A merged fluorescence image is not automatically a single raw image. Explain which channels were combined, what colors were assigned and whether any channel was adjusted separately. If a projection or average combines planes, frames or exposures, identify that operation and its purpose.
When a display combines data from different places or times, make the relationship explicit. A time-lapse or time-averaged image can be scientifically meaningful when identified as such; presenting it as one instantaneous field is misleading.
How major guidance sources differ
| Source | Shared practical rule | Details to watch |
|---|---|---|
| Nature Portfolio | Use minimal, whole-image and consistent processing; do not touch up individual features; disclose tools and methods. | Detailed microscopy and gel/blot requirements; life-science submissions in its journals may require unprocessed original gel and western blot images with the accepted version. |
| Elsevier | Do not enhance, obscure, move, remove or introduce a specific feature; global adjustments must preserve information. | Individual journals can impose more detailed or different requirements. |
| IOP Publishing | Disclose image changes at submission and in the manuscript; process comparable images equally. | Originals may be requested for substantially processed images; edits must not lead the viewer to a different conclusion. |
| Wiley guidance | Use whole-image adjustments, consistent controls and visible composite delineation; disclose nonlinear edits. | Keep originals available on request and follow the particular journal’s instructions. |
| KU Leuven | Prohibits local feature manipulation and sets rules for global adjustments, composites and nonlinear processing. | The institutional guidance was approved December 19, 2017 and the page was last updated February 28, 2024. |
| Council of Science Editors | Limited adjustment and cropping can be acceptable when interpretation is preserved. | Its 2024 recommendations discuss editorial procedures and reproduce principles requiring explicit labeling of grouped images from different gels, fields or exposures. |
These sources establish a common baseline, not a guarantee that every journal will accept the same figure. A journal can require raw files, a particular legend statement or an additional declaration even when the processing itself is scientifically defensible.
Common situations and the defensible approach
| Situation | Defensible approach | Risk to avoid |
|---|---|---|
| A fluorescence image is too dark | Apply a global brightness or contrast adjustment, apply equivalent treatment to comparison images, and retain the original. | Brightening only the cells supporting the hypothesis. |
| A gel contains large empty margins | Crop the margins while retaining relevant bands and controls; keep the full scan. | Cropping away a control or context that changes interpretation. |
| Two nonadjacent lanes are needed in one panel | Show a visible boundary and identify the rearrangement and source lanes in the legend. | Making the lanes appear adjacent in the original gel. |
| Separate microscope fields illustrate one phenomenon | Place them as separate panels or a visibly divided composite and state that they are different fields. | Joining fields into one continuous-looking image. |
| A channel needs a different color map | Use a stated pseudocolor or lookup table and explain channel assignments. | Using color to imply a quantitative relationship that was not measured. |
| A projection improves apparent detail | Describe the projection, averaging or deconvolution and its effect on apparent resolution. | Presenting processed detail as native acquisition resolution. |
If you discover a problem before submission
You made a local edit accidentally
Return to the unprocessed file and rebuild the panel with a global, information-preserving operation. Do not try to hide the mistake by flattening the figure or exporting a new copy without a record.
You no longer have the original
Stop treating the processed file as a reliable substitute. Search the acquisition system and backups for the source and metadata. If the original cannot be recovered, tell the corresponding editor before submission or correction and explain exactly what can and cannot be verified.
The journal asks for raw files
Provide the requested unprocessed files in the format and stage specified by the journal. Nature Portfolio’s life-science instructions, for example, can require original gel and western blot images with the accepted version. That requirement is an example of venue-specific practice, not an automatic rule for every publisher.
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Do not wait for a reviewer to infer the assembly. Contact the editorial office, supply the corrected figure with visible boundaries and provide the complete legend and processing record requested by the journal.
Quick Recap
Final checklist
- Every adjustment preserves information needed to interpret the result.
- No feature was cloned, healed, erased, moved, selectively enhanced or selectively suppressed.
- Comparable images and controls received equivalent processing.
- Crops retain the context, bands and controls needed for the claim.
- Composite boundaries are visible and the legend identifies the source parts.
- Microscopy channel, color-map, nonlinear, projection, averaging and resolution details are recorded where relevant.
- Software, acquisition equipment, settings and processing steps are documented.
- Original files and metadata are preserved and can be supplied if requested.
- The target journal’s current instructions have been checked for raw-image and disclosure requirements.
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