Fig. 4. Apparent sample heterogeneity due to different particle orientations.

a: Image of a negatively stained yeast proteasomes. Classification of the particle images yielded two class averages (insets 1 and 2). Comparison of the two class averages with projections from a resolution-limited model generated from the crystal structure (26) identified the two averages to correspond to a top (inset 3) and a side view (inset 4) of the proteasome. b: Image of negatively stained yeast Sec23p/Sec24p complexes. Classification of the particle images yielded a variety of slightly different class averages. 3D reconstructions of the classes shown in insets 1 to 5, calculated using images of tilted specimens, looked identical, demonstrating that the variations in the class averages are due to slightly different orientations, in which the complexes adsorbed to the carbon film. The scale bars correspond to 50 nm and the inset panels have a side length of 34 nm in a and 32 nm in b. Figure 4b modified and reprinted from (7). Copyright 2001 with permission from the National Academy of Sciences, USA.