(A1, B1, C1, D1) Images from the preBötC of Dbx1+/CreERT2; Rosa26tdTomato mice showing tdTomato in neurons derived from Dbx1-expressing precursors (i.e., Dbx1 neurons). Scale bar in A1 is 20 µm and applies to all panels. C1 and D1 show the same field of view at two different depths (−20 and −10 µm, respectively). (A2, B2, C2, D2) Masks of ROIs obtained by analyzing the corresponding images above. Red ROIs are deemed valid targets by the circularity test, which evaluates somatic shape; blue ROIs that fail the circularity test are rejected. Circularity analyses distinguish somata from auto-fluorescent detritus (A1, A2) as well as contiguous soma-dendrite images (B1, B2) and isolated segments (shafts) of dendrites (C1, C2, D1, D2). Non-somatic auto-fluorescence is rejected because it does not accurately indicate underlying neurons. Dendritic segments are not valid targets because they are difficult to target in the ablation phase of the experiments and their cell bodies are detectable in adjacent focal planes. Often, a cell rejected by the circularity test in one focal plane (e.g., C2, graygray double arrowhead) is validated in the adjacent plane (D2, graygray double arrowhead). When ROIs that pass the circularity test are detected in more than one focal plane, they are validated or rejected according to the priority rule. ROIs from a deeper focal plane (−20 µm) are validated by circularity and thus colored red (C2, circled ROIs). Subsequent detection of overlaying ROIs at the superficial focal plane (−10 µm), which also pass the circularity test, are nonetheless rejected by the priority rule and thus colored blue (D2, circled ROIs). These criteria for target detection are more fully described in ‘Materials and methods’ and Wang et al. (2013).