CURRENTLY UNDER CONSTRUCTION
Read and understand "Information for Authors" for your intended Journal
Every journal has its own figure format, so pay attention to what they are asking for. Not following the instructions can result in published figures with a sub-optimal resolution, shifted colors, shifted brightness/contrast, etc., etc. You've worked so hard to get to a point where you are ready to publish your interesting data, right? So let's make sure to show your data to the world in the best way possible!
In general, most journals will have specific requirements for the following: file format, figure dimensions, color mode and resolution, use of colors, figure annotations.
So let's start by looking at an example and figure out what they are talking about. Below is a set of instructions for figures to be published in one of our favorite journals, Journal of Neuroscience (Information as of Dec. 2021; bold by MK).
Figures Figures must be numbered independently of tables, multimedia, and 3D models and cited in the text. Do not duplicate data by presenting it both in the text and in a figure. A title should be part of the legend and not lettered onto the figure. A legend must be included in the manuscript document after the reference list, and should include enough detail to be intelligible without reference to the text. Figures must be submitted as separate files in TIFF or EPS format and be submitted at the size they are to appear: 1 column (maximum width 8.5 cm), 1.5 columns (maximum width 11.6 cm) or 2 columns (maximum width 17.6 cm). They should be the smallest size that will convey the essential scientific information. Illustrations should be prepared so that they are accessible to our many color-blind readers, so color should only be used if it is necessary to accurately convey the information being presented by the image. Grayscale generally provides a more faithful representation when a single quantity is displayed. Use textures or different line types rather than colors in bar plots or graphs. Figures with red and green are particularly problematic, and should generally be converted to magenta and green. If no suitable combination can be found, consider presenting separate monochrome images for the different color channels. For line drawings that require color, consider redundant coding by adding different textures or line types to the colors. Color figures should be in RGB format and supplied at a minimum of 300 dpi. Monochrome (bitmap) images must be supplied at 1200 dpi. (← MK note: they are talking about graphs and line drawings in raster file format here.) Grayscale must be supplied at a minimum of 300 dpi. For figures in vector-based format, all fonts should be converted to outlines and saved as EPS files to ensure that they are reproduced correctly. Remove top and right borderlines that to not contain measuring metrics from all graph/histogram figure panels (i.e., do not box the panels in). Do not include any two-bar graphs/histograms; instead state those values in the text. All illustrations documenting results must include a bar to indicate the scale. All labels used in a figure should be explained in the legend. The migration of protein molecular weight size markers or nucleic acid size markers must be indicated and labeled appropriately (e.g., “kD”, “nt”, “bp”) on all figure panels showing gel electrophoresis." |
So what does JNeurosci ask for?
| File format | TIFF or EPS |
|---|---|
| Figure dimensions | 1, 1.5, or 2 columns |
| Color mode and resolution | RGB color or grayscale at 300 dpi; monochrome at 1200 dpi |
| Use of colors | Make them color-blind friendly! |
| Figure annotations | scale bars, MW size markers, etc. |
File format: Raster vs. Vector
There are two categories of digital artwork: Raster and Vector images. Raster images are pixel-based files like EM images, and they include formats like TIFF, JPEG, BMP, and PSD. To edit a raster file, you would be using tools like Fiji/ImageJ, Adobe Photoshop, and GIMP. Vector images are object-based files like graphs created in Microsoft Excel and GraphPad Prism, and these images are supported by file formats including EPS, PDF, SVZ, and AI. To work with a vector file, you are probably using Adobe Illustrator, Adobe InDesign, or Inkscape (there are others as well). In general, most journals prefer vector files because they can ensure the highest quality and produce the best results in publication.
Figure dimensions
Figure dimensions are defined by typesetting format for the final PDF (or physical) print, and journal-specific. If your figure is not conformed to the specified dimensions, the journal's typesetter will have to resize it. This often results in loss of resolution in publication.
Color mode and resolution
Most journals will ask you to present your color images in RGB (or more precisely, sRGB) because it is the standard color space for displaying images online. Some print journals might talk about converting your files from RGB to CMYK, which is an ink-based color space and used for printing. Because RGB color space is wider ( i.e., capable of producing many more colors) than CMYK, there will be a color shift when images are converted from RGB to CMYK for print. That is, if a RGB color falls outside of CMYK space, then it will be re-mapped to fit inside the CMYK color space. There are ways to ensure the best possible conversion for CMYK print, and that is a topic for another discussion.
Resolution is set to the number (or sometimes range of numbers) that ensure the best results for publication. For most practical purposes, "dpi" (dots per inch) and "ppi" (pixels per inch) are interchangeable. If you are submitting your figures in a raster format like TIFF, following the resolution specification is critical because it has a direct consequence on how sharp your figures will look in publication. If you are making your figures in a vector format, it is important to follow the specification for halftone (RGB color or grayscale) images.
Your figure may contain a set of grayscale serial EM images, a couple of color 3D reconstruction images in RGB color, and several graphs. This figure would be considered as "combination halftones" because you have both raster and vector images in a single file. Unfortunately, JNeurosci tells you what resolution to use only for halftones (grayscale and color) and monochrome (line drawings), but not the mix of both. So what to do? If you are making your figure as a TIFF (i.e., raster) file, a good rule of thumb is to go for 900 ppi. Some journals will give you a specific number for this type of figure, so RTFM for your intended journal.
Use of colors
If you are using colors to represent specific experimental conditions, make sure they are consistently applied throughout your paper (e.g., LTP = red; control = blue).
Many journals do ask you to make your figures accessible to readers with colorblindness. See this and this for more details.
Figure annotations
Scale bars are required when you use microscope (or other) images as part of your results.
The font size should be big enough for all annotations to be legible. The smallest font should be 7 points or so (some journals will give you specific sizes).
Story-boarding
Decide which piece of data to present in what order: What is your paper about? What is the most logical and coherent order to present your data? Which images/graphs are critical for following and understanding key points of the paper?
Understand constraints of your intended journal: How many figures are you allowed to include in the main paper? Are you allowed to include any supplemental figures? If so, how many and in what format?
Plan each figure: How would you layout each figure? How big will each image/graph be?
Editing raster images non-destructively using Adobe Photoshop
Notes on image manipulation
Assuming Adobe Photoshop version 23.0 (Windows)
Color space
Go to "Edit" > "Color Settings"
In the Color Settings window under "Working Spaces", choose "sRGB IEC61966-2.1" for "RGB".
Click "OK".
Cropping
Adjusting brightness and contrast
Export the final image
Formatting graphs
Putting together a figure using Adobe Illustrator
Assuming Adobe Illustrator version 26.0 (Windows)
Setting up a file
Go to "File" > "New"
In the "new Document" window, enter the following: title (default is "Untitled-1"), width and height (If you don't know the final size, leave it as the Letter size - 8.5 x 11 in), color mode (choose "RGB color", ), raster effects (choose "High (300 ppi)"
A couple of things I find useful:
Rulers: "View" > "Rulers" > "Show Rulers". You can change the unit of the rulers by right clicking on a ruler.
Grids: "View" > "Show Grid". If your Rulers are in inches but the grid is not, you can change the grid here: "Edit" > "Preferences" > "Guides & Grid". Set "Gridline every:" to 1 in and "Subdivisions:" to 8.
Organize your images into layers
Your figures will probably have multiple images and graphs.
Placing images
Editing raster images
Editing graphs (vector images)
Basic drawing
Annotations: text, scale bars,
Checking colors for colorblindness: "View" > "Proof Setup" > "Color Blindness" – There are two options. Check both to see how your figure appears. Can you differentiate different colors in your figure?
Convert text to outlines
But I want to use Microsoft PowerPoint to put together my figures!
Yes, I hear you. You've used PowerPoint all your life and you know it like back of your hand. And yes, you can insert raster images and graphs from Excel as vector images. Just make sure that your raster images satisfy resolution requirements for your journal before inserting into PowerPoint. One thing PowerPoint will not do is to convert text into outlines. So If you have any special characters (like Greek letters and other non-English alphabets) in your figure, they may not be recognized by some readers' computers and/or printers. This can be prevented if you copy and paste those special characters from Character Map in Windows.
