Preparation of Normal Carbon Support Films

• Copper EM Grids, 400mesh (Electron Microscopy Sciences or other EM supply company)
• Formvar powder (Electron Microscopy Sciences or other EM supply company)
• Acetone (research grade)
• Ethyl Alcohol (research grade)
• Ethylene Dichloride (research grade)
• Filter paper (Whatman #1)

• "Scotch" tape
• Glass Petri dishes (100 x 5mm + lids)
• Glass slides (To easily remove the formvar film it is important that the slides are pre-cleaned)
• Coplin jars with lids (Fisher Scientific or other Biological supply company)
• Emitech K950X carbon evaporator (Emitech)

1. Pre-clean copper grids by sonicating them for 10 sec. in acetone followed by 10 sec. in ethyl alcohol. Rinse in deionized water and pour them over filter paper in a dust free environment to dry (Fig. 1).
2. Prepare Formvar solution. Add 0.12g of Formvar powder to 50 ml of ethylene dichloride and mix well on a magnetic stirrer until dissolved (Fig. 2). Pour the solution into a clean Coplin jar (Fig. 3). This solution is hygroscopic but will store well for several months if kept in a brown, sealed bottle.
3. Clean a glass slide with water and detergent. Rinse well with water to remove the detergent then rinse again with de-ionized water and dry with paper towels or KimWipes. Make sure that the slide is free of dust and lint. Blow off any lint with compressed air. Keep the slide in a dry, dust-free environment until use (e.g. on filter paper under an upturned beaker). If the plastic film adheres to the slide and cannot be floated off (Step 5), several different methods of cleaning the slide -- or even NOT cleaning the slide should be tried. Recent attempts (Sept. 2005) have shown that higher-quality, pre-cleaned slides work the best. There have even been reports that a little bit of grease, such as that obtained by rubbing one's finger against the side of the nose, helps to prevent the film from sticking.
4. Coat the clean slide with Formvar by dipping it into the solution. Touch the edge of the slide to filter paper to wick off the excess fluid. Dry the slide upright in a dust-free environment (Fig. 4).
5. The next step is to remove the Formvar film from the slide and place grids on it. Fill the rectangular, black dish with de-ionized water until a meniscus is formed on top of the dish. Then drag the lucite rod (See Fig. 6) across the surface of the water to remove any floating debris. Score the edges of the Formvar film with an acetone-cleaned razor blade (Fig. 5). Thoroughly breathe over the length of the slide to loosen the film, and float the film off onto a clean water surface by slowly immersing the slide into the water at a ~15° angle (Fig. 6). If the film is difficult to see, a bright desk lamp can be used to reflect light off the surface of the water and the film. Place grids, shiny/bright surface down, onto good (uniform, gray color, un-wrinkled, etc.) areas of the film (Fig. 7). Place a small piece of clean, white office paper onto the surface of the grids and film and allow it to soak up water. The grids will show up through the paper when the paper is fully soaked (Fig. 8). Pick up the paper, grids and film with forceps and place on dry filter paper, grid side up, in a covered Petri dish to dry. (Fig. 9).
6. Carbon coat to desired thickness. See the instructions for doing this elsewhere on this web site (Fig. 10).
7. Remove the Formvar film by placing the white paper and grids onto a piece of filter paper saturated with ethylene dichloride. The level of ethylene dichloride should be sufficient to completely soak the backing paper without submersing the tops of the grids. One half hour should be sufficient time to dissolve the Formvar film and not damage the carbon support. Allow the paper to dry.
8. Tape the grids and paper to a glass slide with "Scotch" tape to prevent the grids from being jostled. Place the slide in a covered petri dish. Care must still be used when handling the grids to prevent them from being scattered.

Fig. 6. Floating the film on the surface of water.	Fig. 7. Cleaned grids on the Formvar film.	Fig. 8. White paper placed over grids in order to pick them up.
Fig. 9. Formvar-coated grids drying in petri dish.	Fig. 10. Comparison of carbon coating thickness of normal carbon films and holey or perforated carbon films.

Fig. 1. Grids drying in a petri dish.


Fig. 2. Formvar solution on stirring plate.


Fig. 3. Formvar solution in a Coplin jar.


Fig. 4. Slides drying under beaker.

Fig. 5. Scoring the Formvar film with a razor blade. The film is thicker on the end of the slide and appears milky.