Fig. 1.
LBF analysis of NuMA distribution from 3D images. (a) Flowchart of the imaging processing steps including a graph of the relative density of LBFs of NuMA in 77 nuclei from the three acini depicted in b. The radial distributions of LBFs within each nucleus in the NuMA image are calculated by first generating a segmentation mask from the image of DAPI-stained DNA. The segmentation mask not only defines the extent of each nucleus but is also used to define a set of radial steps within each nuclear volume. The graph shows the mean and standard deviation of the relative density of NuMA bright features extracted by LBF analysis (ordinate) as a function of the relative distance from the perimeter (0.0) to the center (1.0) of the nuclei (abscissa). (b–f) Distribution density of the bright features of NuMA in acinar cells. S1 HMECs were cultured in 3D to induce acinar morphogenesis. Each image corresponds to the application of the different steps of distribution analysis starting from the same original image. (b) Fluorescence micrograph of DAPI-stained nuclei from a single optical section containing three acini [1;2;3]. (c) Fluorescence micrograph of Texas red-immunolabeled NuMA from the optical section corresponding to the DAPI image shown in a. (d) Segmentation mask derived from the DAPI-stained image showing a single slice of individually enumerated nuclei. (e) Composite view of the LBFs (light gray) extracted by the LBF analysis overlaid with the segmentation mask (dark gray). (f) Concentric terraces resulting from the application of the distance transform on the segmentation mask.