- Place a drop of water on a slide. Emulsify a small amount of each of the slant cultures in the same drop of water.
- Ring and code the slide with the diamond marking pencil.
- Stain the slide by the endospore stain:
- Place the slides on a slide rack extended across a beaker of boiling water held on a tripod (an electric burner may be
used instead of a Bunsen burner). An asbestos mat should protect the beaker from the Bunsen flame beneath (fig. 7.2).
- Flood slide with malachite green and allow to steam gently for 5 to 10 minutes. The stain itself should not boil; if it does,
reduce the heat. If the stain appears to be evaporating and drying too rapidly, add a little more. Keep the slide flooded.
- Allow the slide to cool slightly, then use forceps to drain the slide over a sink or staining tray and rinse with water for
about 30 seconds until no more green washes out.
- Counterstain the preparation with safranin for 30 seconds, then rinse again with tap water. Blot or air-dry the slide.
- Examine the smear under the oil-immersion objective and record your observations under Results.
|Figure 7.2 A simple method for applying heat when staining smears, using the endospore stain technique.