Figure 2.

(A) Z-projection images (consisting of all the flattened image slices superimposed at maximum transparency) of nuclei in MRC5 primary fibroblasts. Nucleoli (red, A1), SC35 domains (red, A2–A4), and PML NBs (green, B1–B5) were labeled using primary antibodies directed against nucleolar protein B23, the splicing factor SC35 and PML protein, respectively; and fluorescently tagged secondary antibodies were targeted to the primaries. Telomeres (red, A5) were imaged using FISH labeling with a Cy3-conjugated PNA probe. Each image slice captures 20.8 μm × 20.8 μm. (B) Projection images of the segmentations produced by applying the SCT algorithm to each color channel of the image stacks shown in panel A. Yellow regions represent voxels that have been segmented as both PML NB (green) and red object (nucleoli, SC35 domains, or telomeres). (C) Projection images of the segmentations produced by applying Otsu's algorithm to each color channel of the image stacks shown in panel A.