Many bacteria are surrounded by a slimy layer called a capsule that usually consists of a highly hydrated layer of polysaccharide or in a few cases polypeptide. The capsule can have a number of different functions, helping bacteria escape phagocytic white blood cells, protecting against bacteriophage infection, dehydration or facilitating the adherence of bacteria to surfaces. Staining of the capsule requires that the cells NOT be heat-fixed, since any exposure to heat destroys the capsule.
Students will work individually.
Make a light smear from the milk culture of Klebsiella pneumoniae. Take 1-2 loopfuls and spread it throughout the circle. Do not dilute the culture with water.
Air dry thoroughly. Make certain that there is no evidence of moisture on the slide when you proceed to the next step. DO NOT HEAT FIX!
Flood the slide with 1% crystal violet. Allow the stain to sit for 1 minute.
Rinse very gently with water.
Flood the slide with 20% CuSO4 (copper (II) sulfate) and allow this reagent to sit on the slide for 20 seconds.
Rinse very gently with water.
Air dry; do not blot.
Observe the stain using the 10X, 40X, and oil immersion lenses. Record your observations in the results section of this experiment.
This is a stain of Klebsiella pneumonia. Note the capsule halos around the rod shaped bacteria.
