Oligomers and beaded APFs
The first method of evaluating the sizes of the beads was to copy individual images of isolated beads in both bAPF and sAPF micrographs, classify them according to their sizes, superimpose the images in each category, and then use image averaging to obtain a single image for each size (first two rows of Fig. 2). We identified five sizes of beads. The major difficulty with this method is that the outer perimeter is not clear cut. Although isolated images of the smallest category were difficult to identify in the bAPF micrograph, they were prevalent in some sAPF micrographs.
The second method was to approximate distances between beads of bAPFs. We first identified relatively symmetric bAPF composed of the same size of beads. We used rotational image averaging to obtain symmetric averaged images, and then superimposed images of the same size and shape to obtain a single averaged image. We excluded bAPFs from the analysis if they were relatively asymmetric, had substantial gaps between beads, were composed of different sized beads, contained beads that appear to have merged, and/or had an ambiguous number of beads. The advantage of this approach is that the diameter of the beads can be approximated from the distance between the centers of adjacent beads, which can be measured more precisely than the location of the outer edges of isolated beads. The remaining rows in Fig. 2 are images of assemblies of 2-10 beads. Sizes of the beads are color coded as in the first two rows.