Extended Data Fig. 1: Background subtraction in standard mass photometry (MP) vs dynamic MP.

Zoom on three consecutive raw images from a dynamic MP movie of WT dynamin in contact with an SLB containing a particle diffusing upwards through the image (top row), which is masked by the large signal from the glass surface roughness. In standard MP (left), images are divided by preceding images to remove the large static signal due to surface roughness. This background subtraction relies on stationary binding of small particles to the surface to visualise them. When particles bind to and diffuse on the surface, the background subtraction used in standard MP results in a signal that is a convolution of the particle’s position at t_i-1 and t_i, (blue and red, respectively), which is challenging to reliably detect and quantify. A sliding median filter (right), that is subtracting each image’s temporal median background obtained from a defined window of images around the image of interest, reveals signals of only the particles in the image of interest (t_i, red). For further explanation see ‘Data processing’ in the Methods section. Scale bar = 500 nm. The effect illustrated in this figure was reproducible in all measurements shown in this study (n > 30).