Materials and Reagents

1. Trypsin: working solution is 0.5% trypsin-0.2% EDTA (Life Technologies, Gibco^®, catalog number: 15400-054 )
   
2. NaH₂PO₄ (Kanto, catalog number: 37239-01 )
   
3. Na₂HPO₄ (Chemical, catalog number: 37240-01)
   
4. Alcohol
   
5. Poly/Bed 812 (Electron microscopy sciences, catalog number: 14900 )
   
6. Araldite 502 (Electron microscopy sciences, catalog number: 10900 )
   
7. Dodecenylsuccinic anhydride (DDSA) (TAAB, catalog number: D025 )
   
8. DMP-30 (Electron microscopy sciences, catalog number: 13600 )
   
9. Propylene oxide (Merk, catalog number: S4912527-745 )
   Note: Toxic, need to handle with care and collect after use.
   
10. Syringe
    
11. Dropper
    
12. Grid (200 mesh) (TAAB)
    
13. 0.1 M Phosphate buffer (PB) (see Recipes)
    
14. Paraformaldehyde (4% in PB) (Bionovas, catalog number: AP0130-0500 ) (see Recipes)
    
15. Osimium tetraoxide (1% in PB) (Electron microscopy sciences, catalog number: 51007 ) (see Recipes)
    Note: Toxic, need to handle with care and collect after use.
    
16. Epon (see Recipes)
    
17. Toluidune Blue (Sigma-Aldrich, catalog number: NO202-146-2) (see Recipes)
    
18. Uranyl acetate (Art, catalog number: 8473) (see Recipes)
    
19. Lead citrate (TAAB, catalog number: L003 ) (see Recipes)
    

Equipment

1. Eppendorf tubes
   
2. Centrifuge
   
3. Vacuum drying oven
   
4. Diamond knife (Diatome)
   
5. Ultracut (Leica)
   
6. TEM (HITACHI, model: H-7100 )
   

Procedure