Fig. 5: Segmentation ability of umExM.

a.i Single z-plane confocal image of expanded Thy1-YFP mouse brain tissue after umExM processing, showing pGk13a staining of the membrane. a.ii Single z-plane image showing manual segmentation of the cell body in (a.i). a.iii Overlay of (a.i) and (a.ii). (a.iv) Single z-plane confocal image of the specimen of (a.i), showing GFP signal of the same field of view as in (a.i). (a.v) single z-plane image showing manual segmentation of the cell body from (a.iv). (a.vi) overlay of (a.iv) and (a.v). b As in (a), but for segmenting dendrites. c (left) Single z-plane confocal image of expanded Thy1-YFP mouse brain tissue showing pGk13a staining of the membrane. (c.i) Magnified view of the yellow box on the left. c.ii single z-plane image showing manual segmentation of the myelinated axon in (c.i). c.iii overlay of (c.i) and (c.ii). c.iv Single z-plane confocal image of the specimen of (c.i), showing GFP signal of the same field of view as in (c.i). (c.v) Single z-plane image showing manual segmentation of the myelinated axon in (c.iv). (c.vi) Overlay of (c.iv) and (c.v). d As in (c), but for segmenting unmyelinated axons. (e) Rand score of pGk13a signal-guided segmentation of cell body, dendrites, myelinated axon and unmyelinated axons, using anti-GFP signal-guided segmentation as a “ground truth.” (n = 3 cell bodies and n = 3 dendrites from two fixed brain slices from two mice, and n = 5 myelinated axons and n = 5 unmyelinated axons from two fixed brain slices from two mice). Scale bars: (a.i–vi) 5 μm, (b.i–vi) 5 μm, (c) (left) 2 μm; (i–vi) 0.5 μm, (d) (left) 2 μm; (i–vi) 0.5 μm. Source data are provided as a Source Data file.