Gram Stain Lab - to help study PDF

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Gram Stain Lab Questions

Name__ _______________________________

1. How old can the gram stain culture be? A gram stain culture can be no older than 24-36 hours old to get optimum/reliable results.

2. How long was the decolorizer applied for? The decolorizer (95% Ethanol) was applied for 12 seconds so it can wash the stain out of the thin cell wall bacteria but not the thick cell wall.

3. List the steps of the Gram-staining procedure in order (omit washings), and fill in the color of grampositive and gram-negative cells after each step. Step 1

Chemical Crystal Violet

Gram + Cell Color Purple

Gram – Cell Color Purple

2

Gram’s Iodine

Purple

Purple

3

95% Ethanol

Purple

Colorless

4

Safranin

purple

Pink

4. Which step can you omit without affecting determination of the Gram reaction? The fourth step (safranin) can be omitted without affecting determination of the gram reaction.

5. Suppose you performed a Gram stain on a sample from a pure culture of bacteria and observed a field of red and purple cocci. Adjacent cells were not always the same color. What would you conclude? I would conclude that the bacteria was gram positive; the culture sample was old and the cells lost ability to retain the dye.

6. Suppose you are viewing a Gram-stained field of red rods and purple cocci through the microscope. What do you conclude? I would conclude that the sample has a mixed culture, because red rods are Gram negative and Cocci are Gram positive.

7. Human cells can be stained with crystal violent and safranin, so why can’t human cells be Gramstained? Because human cells do not have a cell wall, so the primary stain would be easily removed by the ethanol. Safranin would bind to remaining structures with a negative charge. Only cells that have a thick cell wall can stain gram positive.

8. What are the three things you want to avoid while doing the gram stain according to the video? According to the video, three mistakes that should be avoided are:  Applying to much bacteria to the bacteria smear  Rubbing slide when blotting dry  Decolorizing for the precise time and immediately rinse...