Most of
the bacteria to be divided into two groups, Gram-positive bacteria and
Gram-negative bacteria.
Gram
staining differentiates bacteria by the chemical and physical properties of
their cell
walls by
detecting peptidoglycan.
Which is
present in a thick layer in gram-positive bacteria.
Discovered
by the Danish scientist and physician Hans
Christian Joachim Gram in
1884.
Fresh
grown or Young culture to be taken.
Because
g+ve bacteria stain not taken.
Staining Procedure:
·
Prepare a thin smear on
glass slide.
·
Heat fix the smear
·
Put one drop of Crystal
violet on the smear wait for 30 seconds.
·
Wash the stain with
purified water.
·
Put one drop of Grams
Iodine wait for 60 seconds.
·
Wash the stain with
purified water.
·
Wash the smear with
Grams Decolonize (95% acetone or Ethanol)for 10-30 seconds and then rinse with
purified water.
·
Put Drop of a Saffron
wait for 30 seconds.
·
Wash the stain with
purified water.
·
Put a drop of oil
immersion on the smear and observe under 100-x objective of the microscope.
Acceptance criteria :
1. Gram positive bacteria (thick layer of
peptidoglycan-90% of cell wall)- stains purple
2. Gram negative bacteria (thin layer of
peptidoglycan-10% of cell wall and high lipid content) –stains
red/pink
