Serial-block face SEM

Serial-Block Face (SBF)-SEM for large-volume tissue and cellular architectures.

Workflow:

  1. Chemically fix tissues/cells with glutaraldehyde & paraformaldehyde mixture
  2. Stain with aqueous osmium tetroxide containing potassium ferrocyanide
  3. Stain with thiocarbohydrazide (TCH) solution
  4. Stain with uranyl acetate solution
  5. Stain with Walton’s lead aspartate
  6. Dehydrate in a series of ethanol dilutions
  7. Infiltrate the specimen into TER-resin
  8. Polymerize at 60°C for 2 days at least
  9. Confirm the target area by conventional ultra-sectioning
  10. Trim to fine tower
  11. Re-embed in the conductive epoxy-resin
  12. Trim and gold sputtering with Quorum K950X Turbo Evaporator
  13. Slice and view using Gatan 3View systems
  14. Image procession by AMIRA2021 machine learning module
  15. Image procession by ilastik (as required)