Exercise 9.7 - TEM Visualization of Microtubules

LEVEL II

Materials

• Coated grid for TEM
  
• 0.1 M ammonium acetate
  
• 5% ethanol saturated uranyl acetate
  
• Transmission electron microscope

Procedure

1. At the conclusion of a 37° C incubation from Exercise 9.6, remove one drop (approximately 20 microliters) of the extract, place it on a collodion or formvar and carbon-coated EM grid, and allow it to remain for about 20-30 seconds. 6

2. Rinse the grid carefully with 0.1 M ammonium acetate, and float the grid upside down on a drop of 5% uranyl acetate for 15 minutes. Rinse the grid with water, blot all excess fluids from the grid, and allow it to air dry.

3. Examine the stained grid in a transmission electron microscope.

4. Draw or photograph your observations -- calculate all appropriate dimensions (length, diameter) of the formed tubules.

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