Serial block face scanning electron microscopy (SBFSEM) can generate large 3D volumes by assembling sequential 2D images acquired in a scanning electron microscope.
SBFSEM incorporates a microtome within the scanning electron microscope chamber. The microtome cuts a thin section from the resin specimen block, an image is collected on the backscatter electron detector, and the procedure is repeated until the required number of images to reconstruct a 3D volume of the sample is generated.
This technique uses room temperature processing techniques with high levels of en bloc heavy metal staining required to generate a sufficiently robust back-scattered electron signal to form an image.