Plastic Embedding of Arabidopsis Stem Sections

Materials and Reagents

1. 30 day-old Arabidopsis stem fragments
   
2. 90% acetone
   
3. 4% glutaraldehyde
   
4. 0.1% Tween 20
   
5. Absolute ethanol
   
6. Historesin standard kit (Reichert-Jung, catalog number: 7O-2218-500)
   Components:
   1. Basic resin liquid (Figure 2A)
      
   2. Activator powder
      
   3. Hardener
      
7. Cresyl violet acetate (Sigma-Aldrich, catalog number: C-5042)
   
8. Preinfiltration solution (see Recipes)
   
9. Infiltration solution (see Recipes)
   
10. Embedding medium (see Recipes)
    
11. Cresyl violet staining medium (see Recipes)
    

Equipment

1. Vacuum pump (Figure 1)
   
2. Vacuum jar (Figure 1)
   
3. Histomold trays (6 x 8 mm) (Leica Microsystems, catalog number: 14702218311)
   
4. Histomold trays (13 x 19 mm) (Leica Microsystems, catalog number: 14702218313)
   
5. Specimen Holder adapter (Leica Microsystems, catalog number: 14702218310)
   
6. Drying oven DO-90 (Raypa)
   
7. Motorized microtome Leica RM2155 (Leica Microsystems)
   
8. Water/ethanol bath HI 1210 (Leica Microsystems)
   
9. Tungsten carbide disposable blades TC-65 (Leica Microsystems, catalog number: 140216266379)
   
10. Glass slides Superfrost ultra plus (Menzel-Glasër, catalog number: J3800AMNZ)
    
11. Surgipath micromount (Leica Microsystems, catalog number: 3801731)
    
12. Coverslip (24 x 60) (Menzel-Glasër, catalog number: BB024060A1)
    
    
    Figure 1. Vacuum jar and vacuum pump used for tissue infiltration
    

Procedure