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A multivesicular body is an endosome containing multiple small vesicles within its lumen. These internal vesicles form when regions of the boundary cell membrane bud inward and undergo separation. This reverses the familiar direction of a transport vesicle budding outward into the cytosol.

Membrane topology explains a useful consequence. A cytosol-facing portion of a membrane protein becomes enclosed inside an internal vesicle; subsequent delivery to a lysosome can expose the whole cargo to degradation. Sorting onto an internal vesicle can therefore remove a signalling receptor from the cytosolic machinery that it previously contacted.

A reconstitution experiment added purified ESCRT-III components to large artificial vesicles and observed formation and detachment of inward buds. This demonstrated a direct membrane-separation activity in a simplified system, rather than merely finding the proteins near cellular multivesicular bodies.^[ref]

Thomas Wollert investigates how proteins reshape cell membranes. In a 2009 study, Wollert and colleagues reconstructed membrane cutting using purified ESCRT-III components and large artificial vesicles. Fluorescence microscopy revealed the formation and detachment of smaller vesicles into their interiors. This reduced a process normally embedded in a complex biological cell to components that could be added and omitted separately.^[ref]

Three ESCRT-III components sufficed for detachment in this system. Additional machinery, including an ATPase, was needed to recycle the protein assembly and sustain further rounds. The distinction is illuminating: the minimum machinery for one cutting event need not be the minimum machinery for repeated operation. The purified system established sufficiency under its experimental conditions, without showing that all processes in living biological cells use exactly that minimal sequence. The paper credits Wollert with preparing artificial vesicles, purifying and labeling proteins, and performing microscopy.^[ref]