Acid Fast Staining (Ziehl-Neelsen Method | P)

Question 1

Give examples of bacteria that are acid-fast

Answer 1

1) Mycobacterium

2) Nocardia

Question 2

Why are acid-fast bacteria called acid-fast?

Answer 2

Because during acid-fast staining, the bacteria retain the primary dye carbol fuchsin (carbolfuchsin) despite decolorization with the powerful solvent acid-alcohol

Question 3

What are the non acid-fast bacteria?

Answer 3

Nearly all other genera of bacteria

Question 4

What is the meaning of AFB?

Answer 4

Acid-fast bacilli

Question 5

What is the characteristic of AFB?

Answer 5

They have mycolic acid in their cell walls

Question 6

What is the purpose of mycolic acid?

Answer 6

To prevent the acid-alcohol from decolorizing the protoplasm

Question 7

What type of stain is acid-fast stain?

Answer 7

It is a differential stain

Question 8

What is the procedure of Ziehl-Neelsen staining?

Answer 8

1) Apply primary stain of carbolfuchsin (for 30 secs)
2) Heat fix cells to slide (by the use of flame)
3) Decolorize with acid alcohol (for 15 - 20 secs)
4) Apply counterstain of methylene blue (for 30 secs) then rinse excess stain

Question 9

What is the characteristic (in terms of shape) of AFB?

Answer 9

Rod-shaped (bacilli)

Question 10

Explain the result for Mycobacterium and non-Mycobacterium in acid-fast staining

Answer 10

1) Mycobacterium and non-Mycobacterium: all bacteria are transparent prior to staining | resulting color: both transparent
2) Myco and non-Myco: steam is used to force the carbol fuchsin (carbolfuchsin) into all the bacteria | resulting color: both purple
3) Myco and non-Myco: decolorization with acid removes the carbol fuchsin (carbolfuchsin) from the bacteria that do not have mycolic acid | resulting color: purple and pink
4) Myco and non-Myco: methylene blue is the counterstain | resulting color: purple and blue

Question 11

What is the resulting color for AFB?

Answer 11

These bacteria retains the primary dye, hence, these results to color red or pink

Question 12

What is the resulting color for non-AFB?

Answer 12

Non-AFB takes up the counterstain, hence, these results to color blue or green

Question 13

Explain and provide the reading and interpretation of results for AFB

Answer 13

0: No AFB seen in 300 fields
+^N: 1 - 9 AFB seen in 100 fields
1+: 10 - 99 AFB seen in 100 fields
2+: 1 - 10 AFB seen per field in at least 50 fields
3+: > 10 AFB seen per field in at least 20 fields