Difference: PlasticCoatedGrid (7 vs. 8)

Revision 818 Feb 2011 - Main.BillRice

 
META TOPICPARENT name="PPNegativeStaining"
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Principles & Protocols

Protocols - Plastic Coated Grids

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You will need:

  • Pyrex crystallizing dish
  • Metal rod
Changed:
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  • Amyl acetate
>
>
  • 1% Nitrocellulose dissolved in amyl acetate
 
  • Tweezers
  • 400 mesh square Cu grids
  • Filter paper
  • Village Voice newspaper
  • Pennies
  • Tissue paper

Procedure:

Changed:
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<
  • Fill a "Pyrex crystallizing dish" to the brim with water
  • Dip a metal rod in amyl acetate
>
>
  • 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
Changed:
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  • Throw the first collodion layer away
  • Repeat amyl acetate drop on the water and allow evaporation
  • Place grids on plastic
  • Cut a relatively ink free piece from the "Village Voice"
>
>
  • 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

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