a, An X-shaped fiducial (within the red square) in ion-beam view is used for the registration of FIB slicing. b, An X-shaped fiducial (within the yellow square) in electron-beam view is used for registration of electron imaging. The round cross-section of the perpendicularly drilled hole (within the orange square) is used for the fine registration of secondary-electron images. c, A secondary-electron image used for data acquisition, showing an ion-milled hole (within the orange square) used for fine registration. d, Monitoring of slice thickness: we measured the distance between the milling surface of the nth slice and the milling surface of the first slice (total slice thickness d) from FIB images, and then plotted d versus (n − 1). This reveals a linear relationship with a slope of 2.96 ± 0.01, which is the averaged slice thickness (in nm).