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Western blot and SDS-PAGE molecular weight calculator

Q: How do you calculate the molecular weight of a protein from a gel?

Plot log₁₀ of the ladder sizes against their migration distances, fit a straight line, and read your band’s size from its distance. This tool does it from clicks on the image.

Upload a photo of your gel or blot, click the ladder bands (their sizes fill in from the ladder you choose), then click your sample bands. The calculator fits log₁₀(molecular weight) against migration distance and reports each band’s estimated size in kDa, with R², an annotated image and a CSV. It also works with distances typed in by hand.

What it does: Estimates protein sizes in kDa from a gel or western blot image: click the ladder and sample bands and it fits a log-linear standard curve with R².
Formula: $\begin{aligned} &\log_{10}(\mathrm{MW}) = a + b \times \text{migration distance} \\ &\text{(least squares on the ladder bands)} \end{aligned}$
Example: With the 70 kDa band at 210 px and the 55 kDa band at 262 px, a sample band at 236 px is about 62 kDa.

Last updated 8 November 2026

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At a glance

Western blot MW calculator at a glance
Input	An image of your blot; click the ladder and sample bands
Ladders	PageRuler, PageRuler Plus, Precision Plus
Model	$\log_{10}(\mathrm{MW})$ linear in migration distance, fitted with R²
Flags	Bands outside the ladder range (extrapolated)
Output	Annotated image and calibration plot (PNG), CSV
Runs in	Your web browser; nothing is uploaded
Price	Free, no account needed

What it does

How to use the western blot MW calculator

Upload the imageDrop a PNG or JPEG of the gel or blot; it stays in your browser.
Mark the ladderChoose your ladder and click its bands from top to bottom; correct any size in the table.
Mark your bandsSwitch to sample bands and click each band of interest; read the estimated sizes and export.

How it works

$\log_{10}(\mathrm{MW}) = a + b \times \text{distance}$

Over a gel’s resolving range, a protein’s migration distance is approximately linear in the logarithm of its molecular weight. The ladder bands give the calibration line by least squares; each unknown band’s distance is read off that line. Distances are measured from the top of the image, which gives the same result as using relative mobility ( $R_\mathrm{f}$ ) because $R_\mathrm{f}$ is a linear rescaling of distance.

Worked example

If the 70 kDa ladder band runs 210 px from the top and the 55 kDa band 262 px, a sample band at 236 px sits between them, at about 62 kDa, assuming the calibration line fits well ( $R^2 \ge 0.98$ ).

Frequently asked questions

How do you calculate the molecular weight of a protein from a gel?

Plot $\log_{10}$ of the ladder sizes against their migration distances, fit a straight line, and read your band’s size from its distance. This tool does it from clicks on the image.

Why does my protein run at a different size than predicted?

Post-translational modifications (glycosylation, phosphorylation), unusual charge or structure, incomplete denaturation and SDS binding differences can all shift apparent molecular weight by 10 % or more.

Which ladder bands should I use?

Those that bracket your protein and are well resolved. The relationship is linear only over the gel’s resolving range, so drop bands compressed at the top or bottom if R² is low.

Is the image uploaded?

No. It is processed in your browser only.

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