Fig. 2: A semi-automated correlative imaging method to identify cell fate decisions in cerebral organoids. | Nature Cell Biology

Fig. 2: A semi-automated correlative imaging method to identify cell fate decisions in cerebral organoids.

From: A cell fate decision map reveals abundant direct neurogenesis bypassing intermediate progenitors in the human developing neocortex

Fig. 2: A semi-automated correlative imaging method to identify cell fate decisions in cerebral organoids.

a, Schematic representation of correlative microscopy pipeline. b, Step-by-step protocol for semi-automated correlative microscopy. (1) bRG cells are live imaged at 20X for 48 hours. (2) 4X brightfield images containing the video coordinates are assembled. (3) Organoid slices are fixed, immunostained for SOX2, EOMES and NEUN, and imaged. (4) Images are automatically segmented to outline slices from live and fixed samples. (5) Slice contours are automatically paired based on shape and area and (6) aligned (including a horizontal flip if needed). (7) Video fields of view are automatically annotated on the immunostaining images. (8) Regions of interest are re-imaged at higher resolution (×40) and cells from live and fixed samples are manually matched. c, Live/fixed correlative analysis of a dividing bRG cell generating a self-renewing bRG daughter and a differentiating IP daughter. d, Live/fixed correlative analysis of a dividing IP cell generating two neuronal daughters. e, Live/fixed correlative analysis of a migrating neuron. All images are representative examples of experiments performed at least three times independently (N = 1,101 bRG cells).

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