Lab 4 : Structural Staining

Question 1

Acid Fast staining - Cold method procedure:

Answer 1

1. Begin with heat-fixed smear
2. Cover with Kinyoun’s Carbol Fuchsin for 5 min
3. Rinse with DH2O
4. Decolorize with Acid Alcohol
5. Rinse with DH2O
6. Counterstain with Methylene Blue for 1 min
7. Rinse with DH2O
8. Blot dry

Question 2

Acid Fast stain result: RED

Answer 2

Acid Fast

Question 3

Acid Fast stain result: BLUE

Answer 3

Non-Acid Fast

Question 4

Endospore Staining procedure:

Answer 4

1. Begin with heat-fixed smear
2. Set up water bath
3. When water is boiling place a piece of absorbent paper on top of the smear
4. Place smear over the water bath and cover it with Malachite Green for at least 7 min. Keep adding Malachite Green and make sure the paper is always moist
5. After the 7 min remove smear from water bath and remove the paper
6. Rinse with DH2O
7. Counterstain with Safranin for 1 min
8. Rinse with DH2O
9. Blot dry with bibulous paper

Question 5

Which two genera of bacteria produce endospores?

Answer 5

Clostridium and Bacillus

Question 6

Endospores

Answer 6

Resistant structures made through a process known as sporulation in response to extreme environmental conditions

Question 7

Capsular Staining procedure:

Answer 7

DO NOT HEAT FIX
1. Place a drop of Nigrosin on a slide
2. Transfer a loopful of bacteria to the drop of stain and mix
3. Spread mixture with another slide
4. Allow to air dry
5. Cover smear with Safranin for 1 min
6. Rinse with DH2O
7. Blot dry