Figure 5.

Midbody and Membrane Markers Are Correctly Localized in Isolated Intact Midbodies

(A and B) Isolated midbodies were stained with α-tubulin, MKLP-1, a marker for midbodies (A), and FM4-64, a membrane marker (B). Scale bar, 5 μm.

(C) Representative electron micrograph shows an isolated midbody with intact membrane structures. Black arrows show vesicles inside, and red arrow shows plasma membrane that surrounds the midbody. Scale bar, 500 nm.

(D) Schematic of the experiments that we conducted to exclude from our LC-MS analysis membrane impurities that copurify with midbodies. We obtained and compared the profiles of midbodies versus cytokinesis (MB-CYT) and midbody versus purified lysate (MB-LYS). The hits from MB-CYT and MB-LYS profiling were compared. MB-CYT hits were checked in MB-LYS profiling. Lipids that were unchanged in MB-LYS profiling (i.e., found in lysate from asynchronous cells in equal amounts) were eliminated, as they are most likely impurities that copurified in both samples. Membrane structures, microtubules, and DNA are cartooned in gray, green, and red, respectively.