A total of 676 cells co-expressing GFP-Atg8 and mHtt(72Q)-mCherry were examined for co-localization. Of these, 294 contained mHtt IBs, and 101 contained both mHtt IBs and autophagosomes. We measured the average apparent diameter of the IBs and autophagosomes. Based on the apparent area of the IB and autophagosome compared with the available cytoplasmic cross-sectional area of the cell, we estimated that they will overlap by chance ∼5–8% of the time. Because more than 90% of autophagosomes are below the limit of resolution, and the IB is often below the limit of resolution, apparent overlap does not signify contact. Of 101 cells containing both IBs and autophagosomes, there were two instances of partial overlap, below the threshold for significance. (A) A maximum projection of a typical cell with both a GFP-Atg8–labeled vesicle (green) and an mHtt(72Q)-mCherry (magenta) IB is shown. The IB and autophagosome are in different axial planes as well as in different locations in the X–Y plane. (B) Autophagosomes containing the GFP-Atg8 fusion protein are competent to fuse with the vacuolar membrane in cells expressing mHtt-mCherry. The scale bar represents 2 μm in (A) and (B).