METHODS: Standard Plastic Embedding Protocol

Stock Solutions:
0.1 M phosphate buffer, pH 7.4 (PB)
100% Acetone
100% anhydrous MeOH and 75%, 85%, 95% MeOH in dH2O
DMP30
Eponate resin
DDSA

Mixing Active Resin: Mix resin fresh or thaw from freezer. Work quickly to prevent components from hydrating. Wear gloves as these components may cause contact dermatitis and the synthesis of certain components involves potential carcinogens. Prepare under a hood.

• Mix 2 volumes DDSA and 1 volume Medcast gently by hand with a spatula until the solution is clear (2-5 minutes)
• Add 0.6 ml DMP30 per 30 ml resin; Mix gently as before until clear (2-5 minutes). Solution will darken as the activated epoxide begins to bind oxygen.
• Use immediately or store in 30-60 cc syringe in the freezer. Expel all air before freezing and plug syringe with a wooden dowel. Do not cap with a needle or plastic cover as they will leak air.

Dehydration and Infiltration Schedule: All volumes 5-10 ml unless noted otherwise. This procedure is designed for tissue samples < 1 mm thickness, presumed to be in fixative.

• 10 m PB
• 10 m 75% MeOH
• 10 m 85% MeOH
• 10 m 95% MeOH
• 10 m 100% MeOH
• 10 m 100% MeOH
• 10 m 100% Acetone
• 10 m 100% Acetone
• 60 m 50% resin in acetone
• 4-20 h 75% resin in acetone
• 60 m 100% resin
• Embed samples in fresh resin in molds and cure at 60-65 deg C overnight

Disclaimer: As conditions of use are outside our control, we make no warranties, express or implied, and assume no liability in connection with use of this protocol.

© 2000 Robert E. Marc
This protocol may be copied and distributed for educational and non-profit
purposes as long as acknowledgement of our copyright is included
in every instance of use. It may not be sold or distributed for commercial gain.