Basic Microbiology Group 1 :
Alifia N Fadila (2001541831)
Amarsha Darnidita P (2001547103)
Clarice Angelica (2001542752)
Jessica (2001554941)
Odilia Odny Pangestu (2001541314)
Shierly Chandra (2001559356)
Vania Clarensia (2001584313)
Question: In which phase gram staining should be conducted? Why?
Gram Staining is an important diagnostic staining technique for bacteria. It permits ready
differentiation of major categories based upon the color reaction of the cells: gram positive,
which are stained purple, and gram-negative, which are stained red. This difference in staining
quality is due to structural variations found in the cell walls of bacteria.
In the first step, crystal violet stains cells in a smear all the same purple color. The second
and key differentiating step is the mordant, Gram’s iodine. The mordant causes the dye to form
large crystals that get trapped by the meshwork of the cell wall. Because this layer in gram-
positive cells is thicker, the entrapment of the dye is far more extensive in them than in gram-
negative cells. Application of alcohol in the third step dissolves lipids in the outer membrane of
the gram-negative cells, which removes the dye from them. By contrast, the crystals of dye
tightly embedded in the gram-positive bacteria are relatively inaccessible and resistant to
removal. Because gram-negative bacteria are colorless after decolorization, their presence is
demonstrated by applying the counterstain safranin in the final step.
Gram staining is a procedure which depends upon the cell wall composition of bacteria.
Thus bacteria in log phase have the well developed cell wall which gives the correct reaction
leading to the identification of bacteria as gram positive and gram negative. Also in the log
phase, the shape and conformation of bacteria, especially the cell walls are more stable.
In older cells, the cell walls having become weak may get decolourized during gram
staining and always almost appear gram negative which may be incorrect. So to get the correct
results it’s required to have cells with intact cell walls. Which is why log culture cells are the best
to use. Cells from old cultures may stain Gram negative even if the bacteria are Gram positive.
Cells in the exponential phase of growth are the healthiest and most uniform, which explains
why most experiments utilize
cells from this phase.
Alifia N Fadila (2001541831)
Amarsha Darnidita P (2001547103)
Clarice Angelica (2001542752)
Jessica (2001554941)
Odilia Odny Pangestu (2001541314)
Shierly Chandra (2001559356)
Vania Clarensia (2001584313)
Question: In which phase gram staining should be conducted? Why?
Gram Staining is an important diagnostic staining technique for bacteria. It permits ready
differentiation of major categories based upon the color reaction of the cells: gram positive,
which are stained purple, and gram-negative, which are stained red. This difference in staining
quality is due to structural variations found in the cell walls of bacteria.
In the first step, crystal violet stains cells in a smear all the same purple color. The second
and key differentiating step is the mordant, Gram’s iodine. The mordant causes the dye to form
large crystals that get trapped by the meshwork of the cell wall. Because this layer in gram-
positive cells is thicker, the entrapment of the dye is far more extensive in them than in gram-
negative cells. Application of alcohol in the third step dissolves lipids in the outer membrane of
the gram-negative cells, which removes the dye from them. By contrast, the crystals of dye
tightly embedded in the gram-positive bacteria are relatively inaccessible and resistant to
removal. Because gram-negative bacteria are colorless after decolorization, their presence is
demonstrated by applying the counterstain safranin in the final step.
Gram staining is a procedure which depends upon the cell wall composition of bacteria.
Thus bacteria in log phase have the well developed cell wall which gives the correct reaction
leading to the identification of bacteria as gram positive and gram negative. Also in the log
phase, the shape and conformation of bacteria, especially the cell walls are more stable.
In older cells, the cell walls having become weak may get decolourized during gram
staining and always almost appear gram negative which may be incorrect. So to get the correct
results it’s required to have cells with intact cell walls. Which is why log culture cells are the best
to use. Cells from old cultures may stain Gram negative even if the bacteria are Gram positive.
Cells in the exponential phase of growth are the healthiest and most uniform, which explains
why most experiments utilize
cells from this phase.
