THE VIRTUAL EDGE: Lab 3 Bacterial Staining Techniques II

Gram Stain: Background & Introduction

Gram +

Gram-positive cell walls have a thick peptidoglycan layer beyond the plasma membrane. Characteristic polymers called teichoic and lipoteichoic acids stick out above the peptidoglycan and it is because of their negative charge that the cell wall is overall negative. These acids are also very important in the body’s ability to recognize foreign bacteria. Gram-positive cell walls stain blue/purple with the Gram stain.

gram+

Gram -

Gram-negative cell walls are more complex. They have a thin peptidoglycan layer and an outer membrane (lipopolysaccharide) beyond the plasma membrane. The space between the plasma membrane and the outer membrane is called the periplasm. The outer leaflet of the outer membrane is composed mainly of a molecule called lipopolysaccharide (LPS). LPS is an endotoxin that is important in triggering the body’s immune response. Gram-negative cells will stain pink with the Gram stain.

Gram-

Procedure	Reagent	Gram Positive	Gram Negative
Gram Stain		Cell Color
Fixed cells on slide		COLORLESS	COLORLESS
Primary stain	Crystal Violet	PURPLE	PURPLE
Mordant	Iodine	PURPLE	PURPLE
Decolorizer	Alcohol	PURPLE	COLORLESS
Counterstain	Safranin	PURPLE	RED