Serial-Block Face (SBF)-SEM for large-volume tissue and cellular architectures.
Workflow:
- Chemically fix tissues/cells with glutaraldehyde & paraformaldehyde mixture
- Stain with aqueous osmium tetroxide containing potassium ferrocyanide
- Stain with thiocarbohydrazide (TCH) solution
- Stain with uranyl acetate solution
- Stain with Walton’s lead aspartate
- Dehydrate in a series of ethanol dilutions
- Infiltrate the specimen into TER-resin
- Polymerize at 60°C for 2 days at least
- Confirm the target area by conventional ultra-sectioning
- Trim to fine tower
- Re-embed in the conductive epoxy-resin
- Trim and gold sputtering with Quorum K950X Turbo Evaporator
- Slice and view using Gatan 3View systems
- Image procession by AMIRA2021 machine learning module
- Image procession by ilastik (as required)