SPECIAL STAINS PART 2

Question 1

Why is gram staining performed

Answer 1

It’s a differential staining technique that represents
an important initial step in the characterization and classification of bacteria using a light microscope

Question 2

Who discovered gram staining

Answer 2

Hans Christian Gram in1884

Question 3

Why is gram staining performed in histology

Answer 3

Bacteria are the most difficult organisms to detect in H&E-stained sections

Question 4

What histologic changes are suggestive of a bacterial infection

Answer 4

consistent pattern of inflammation

Question 5

What is done to a tissue when histological changes are suggestive of a bacterial infection

Answer 5

Gram-stained section of the tissue
can then be helpful to determine if bacteria are indeed present.

Question 6

What is the difference between gram negative and gram positive bacteria based on peptidoglycan layer

Answer 6

It is thicker in gram positive bacteria

Question 7

What is the difference between gram negative and gram positive bacteria based Lipopolysaccharide (a lipid layer external to peptidoglycan)

Answer 7

It is only present in gram negative bacterias

Question 8

explain the general principle of gram staining

Answer 8

1. Initial staining with crystal violet. This is a basic dye that similarly stains both
   Gram-positive and Gram-negative organisms.
2. Treatment with an iodine and potassium iodide solution (mordant) that serves
   to fix the crystal violet stain. Both Gram-positive and Gram-negative
   organisms remain purple after this step.
3. Decolorization with a mixture of alcohol and acetone. This is the differential
   step: Gram-negative bacteria become destained, while Gram-positive bacteria
   retain the color of the crystal violet-iodine complex.
4. Counterstaining with fuchsin (red) or safranin (pink). Because the Grampositive bacteria are already stained purple, they are not affected by the
   counterstain. However, the now colorless Gram-negative bacteria do become
   stained

Question 9

Explain the influence of the bacteria cell wall and difference in staining

Answer 9

The difference in staining between Gram-positive and Gram-negative bacteria is
based on their ability to retain the color of the violet stain used during the reaction.
The lipopolysaccharide layer of Gram-negative bacteria holds the key to this. It is
disrupted in the decolorization step. This allows the original crystal violet stain to
leach out, and the basic fuchsin or safranin counterstain to be taken up.
The thicker peptidoglycan layer of Gram-positive bacteria, on the other hand, allows
the crystal violet stain to be retained.