Gel and western blot lane labeller

$$\text{lane width}=(x_1-x_0)/n$$$$\begin{array}{rl}& \text{centre of lane }i(i=0\dots n-1)\\ & =x_0+(i+\frac{1}{2})\times \text{lane width}\end{array}$$$$\begin{array}{rl}& \text{first guess for ladder band }M:\\ & y=y_{\mathrm{top}}+\frac{\ln M_{\mathrm{top}}-\ln M}{\ln M_{\mathrm{top}}-\ln M_{\mathrm{bottom}}}\\ & \times (y_{\mathrm{bottom}}-y_{\mathrm{top}})\end{array}$$$$\begin{array}{rl}& \text{height an angled label needs}\\ & =\text{text width}\times \sin \theta +\text{font size}\times \cos \theta \end{array}$$

Lanes. Wells are cast at equal spacing, so the lanes start evenly divided over the area the current lanes cover (the whole width for a new image); drag each green marker onto the middle of its lane where a gel ran unevenly or was cut. The labels are centred on, or for angled labels start at, the lane centres, above the image.

The marker column. In SDS-PAGE the logarithm of a protein’s molecular weight falls about linearly with the distance it migrates, which Weber and Osborn established with 40 proteins of known size (Weber & Osborn 1969); DNA fragments in agarose behave similarly over the middle of a gel’s range. The tool uses this only for a first guess: the top and bottom band of the chosen ladder go near the top and bottom of the image and the bands between are spaced on that log scale. Then drag each orange mark onto the real band in the marker lane, because gradient gels, prestained proteins and the ends of the range depart from the straight line. To read the size of an unknown band from the ladder, use the western blot molecular weight calculator. Prestained ladders such as PageRuler give approximate sizes: the manufacturer notes that their apparent molecular weight varies with the gel and buffer system and from lot to lot.

Rotating and cropping. Rotation turns the image about its centre into a canvas just large enough to hold it (width $W\cos \theta +H\sin \theta$, height $W\sin \theta +H\cos \theta$), with the corners filled white or black, and then the crop removes the number of pixels you set from each side. Rotation by angles other than 90° resamples the image. Image-integrity guidance accepts straightening, cropping and labelling, but adjustments must be applied to the whole image, and blots should not be cut and spliced without marking the joins (Rossner & Yamada 2004); keep the unprocessed original.

Output. The PNG has the image at its own pixel size with margins added for the labels and marker column, and keeps the DPI the original file recorded. The SVG embeds the processed image and writes labels, marks and unit as text in Arial, so they can be edited in Illustrator, Inkscape or the LabCanvas figure editor.

Sources: Weber K, Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem 1969;244:4406–4412, doi:10.1016/S0021-9258(18)94333-4. Rossner M, Yamada KM. What’s in a picture? The temptation of image manipulation. J Cell Biol 2004;166:11–15, doi:10.1083/jcb.200406019. Ladder sizes: the manufacturers’ product pages linked in the table.

A western blot image 900 × 520 pixels: the ladder in lane 1, then a control, two siRNAs and a time course (0, 2, 6 and 24 h). The lanes run from $x=50$ to $x=850$.

1. Lanes. 8 lanes over 800 pixels are 100 pixels wide; their centres are at $50+(i+\frac{1}{2})\times 100$ $=100,200,300\dots 800$ pixels.
2. Labels. Type M, Ctrl, siRNA 1, siRNA 2, 0 h, 2 h, 6 h, 24 h and choose 45°: each label starts at its lane centre and rises to the right, and the figure gains just enough space above the blot for the longest one.
3. Marker. Choose PageRuler Plus Prestained. With its 250 kDa band at $y=60$ and its 10 kDa band at $y=470$, the bands between are spaced as below: 55 kDa at $60+(\ln 250-\ln 55)$ $÷(\ln 250-\ln 10)\times 410=252.9$ pixels. On the example blot these sit exactly on the bands; on a real gel, drag each mark onto its band.
4. Export. Download the PNG for a slide or the SVG to keep the labels editable in a figure.

For a DNA gel, GeneRuler 1 kb from 10 000 bp at the top to 250 bp at the bottom works the same way; its brighter reference bands at 6000, 3000 and 1000 bp help you match the marks to the right bands.