Gram Staining

Question 1

The Gram stain is an example of what type of stain?

Answer 1

Differential Stain

Question 2

What is a deferential stain?

Answer 2

Differentiating cell types, which will be stained different colors as a result of the procedure.

Question 3

What is the procedure of a gram stain and its purpose? What would be the result of looking under the microscope at each step?

Answer 3

*After we do the simple heat-fix smear
1. Cover smear with crystal violet 30-60 sec, then wash; everything purple!
2. Rinse with iodine, then cover with iodine for 30-60 sec, then wash; everything still purple!
3. Decolorize gram stain with 95% ethanol 20-30 sec, then wash IMMEDIATELY; Gram+ still purple! Gram- is clear.
4.Counterstain with Saffranin 30-60 sec, then wash and blot+let dry; Gram+ still purple! Gram- now pink/red.

Question 4

Why is the decolorizing step time sensitive in the gram staining procedure?

Answer 4

Because it’s possible to over decolorize smears so that gram+ bacteria lose the stain complex, or to under decolorize smears so that the Gram- bacteria don’t lose the stain complex

Question 5

Why can we restain with Saffrinin and still have differential staining?

Answer 5

Because gram+ is already purple, even though it’s stain pink, the purple overpowers it.