Example of an XYZ-drift and a resulting kymograph that is unsuitable for CSC speed analysis
(A) Sum projection of all images from a time-lapse image series with XYZ-drift without StackReg drift correction.
CSC tracks are blurry and not clearly distinguishable.
(B–E) Unsuccessful XYZ-drift correction with StackReg. Time course at 0 (B), 1 (C), 2 (D) and 3 min (E) after XY-drift correction with StackReg reveals substantial drift in the Z-direction. At the beginning of the movie, plasma membrane-bound CESAs are clear (B) but at the end of the movie the focal plane moves inside of the cell where large, bright CESA-containing Golgi stacks are more prevalent (E).
(F) Sum projection of all images from the StackReg-corrected time-lapse image series (B-E) with XYZ-drift. CSC tracks can be seen more clearly than before correction (A) but are still blurry and mixed between each other due to Z-drift. Scale bars A–F: 20 μm.
(G) An example of a kymograph that is unsuitable for analysis. CSC tracks are smeared and not clearly visible; therefore, a speed analysis cannot be performed. Scale bar: 2 μm.