Figure 4. Example application of CL pointer alignment.

Thin section of rat pancreas with post-embedding immunolabeling for insulin (Alexa594, orange). (a) FM, boxed areas indicate regions of interest selected for SEM, based solely on FM appearance in terms of either isolated or dense fluorescence. Data from dashed boxed areas is shown in Supplementary Fig 1. (b) Magnified view of solid boxed area in (a), followed by (c) scanning EM and (d) overlay. Arrows indicate isolated fluorescence features and the corresponding areas in SEM determined after generating the overlay. In the overlay, fluorescence appears well centered on distinct ~100–200 nm granules, while these granules do not a priori stand out in the SEM image. See Supplementary Fig. 1 for further data. Scale bar is 20 μm in (a), 3 μm in (b–d).