Chunk #164 — STAR★Methods — Quantification and Statistical Analysis — Image analysis for astrocyte markers — Analysis of Allen Mouse Brain Atlas in situ images
In order to independently validate our findings about the spatial distribution of astrocytes (Figure S5C), we analyzed in situ hybridization images of marker genes obtained from the Allen Mouse Brain Atlas (http://mouse.brain-map.org) (Lein et al., 2007). First, we downloaded sagittal section images of the following marker genes: Islr, Gdf10, Agt, Mfge8. For overlay of these images and we used similar strategy as described above fro RNAscope. Reference points (16 points) were manually registered in each image. We used the “Expression” images and the following steps. (1) Transform to the common coordinates (MATLAB, “fitgeotrans” function). (2) Transform image from RGB to grayscale (“rgb2gray”). (3) Binarize image using fixed threshold of 100, followed by fill holes (“imfill”). (4) Remove object larger than 2000 pixels and smaller than 100 pixels (“bwareaopen”). (5) Display objects overlay.
