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Principles & Protocols

Protocols - Plastic Coated Grids

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You will need:

• Pyrex crystallizing dish
• Metal rod (or glass pipette)
• p20 pipetman
• 1% Nitrocellulose dissolved in amyl acetate
• Tweezers
• 400 mesh square Cu grids
• Filter paper
• Village Voice newspaper (or blank newsprint paper)
• Pennies
• Tissue paper

Procedure:

• Fill a "Pyrex crystallizing dish" to the brim with water (use a small dish)
• Dip a metal rod in amyl acetate (Better: pipette 20 ul of amyl acetate solution and drop onto water)
• Allow a drop to fall onto the water
• Wait for the amyl acetate to evaporate leaving behind a collodion plastic layer
• Throw the first collodion layer away
• Repeat amyl acetate drop on the water and allow evaporation (few minutes, should see a thin plastic film when done)
• Place grids on plastic (shiny side down)
• Cut a relatively ink free piece from the "Village Voice" (or use blank newsprint -- stocked at NYSBC)
• Place the newspaper on top of the grids
• Gently push the corners of the newspaper into the water with tweezers
• Wait for the newspaper to become saturated with water and all grids to be visible through the paper
• Use a pair of tweezers to bring in the extra plastic film around the newspaper
• Vertically lift the newspaper out of the water
• Place newspaper grid side up on filter paper to dry
• Secure the corners of the newspaper with pennies to keep them from curling
• Carbon Coat the grids once the newspaper is dry

Visit Cryo EM Web site at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005