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Gram stain (Gram staining or Gram's method), is a method of staining used to classify bacterial species into two large groups: gram-positive bacteria and gram-negative bacteria. It may also be used to diagnose a fungal infection. [1] The name comes from the Danish bacteriologist Hans Christian Gram, who developed the technique in 1884. [2]
Violet-stained gram-positive cocci and pink-stained gram-negative bacilli. In bacteriology, gram-positive bacteria are bacteria that give a positive result in the Gram stain test, which is traditionally used to quickly classify bacteria into two broad categories according to their type of cell wall.
Gram Primary stain: Crystal violet applied to film then treated with iodine (mordant), alcohol (decolourizer) and counter stained with safranin Characterizes bacteria in one of two groups, Gram positive or Gram negative Gram positive appears purple in color Gram negative appears pink in color 4 Acid fast (Ziehl-Neelsen technique)
Gram-positive bacteria have a thick peptidoglycan layer in their cell wall, which retains the crystal violet during Gram staining, resulting in a purple color. Gram-negative bacteria have a thin peptidoglycan layer which does not retain the crystal violet, so when safranin is added during the process, they stain red.
The Gram stain, developed in 1884 by Hans Christian Gram, characterises bacteria based on the structural characteristics of their cell walls. [175] [75] The thick layers of peptidoglycan in the "Gram-positive" cell wall stain purple, while the thin "Gram-negative" cell wall appears pink. [175]
In Gram-negative bacteria an outer membrane is also included. [1] This envelope is not present in the Mollicutes where the cell wall is absent. Bacterial cell envelopes fall into two major categories: a Gram-positive type which stains purple during Gram staining and a Gram-negative type which
Gram-positive cell walls are thick and the peptidoglycan (also known as murein) layer constitutes almost 95% of the cell wall in some Gram-positive bacteria and as little as 5-10% of the cell wall in Gram-negative bacteria. The peptidoglycan layer takes up the crystal violet dye and stains purple in the Gram stain.
Gram-negative bacteria will stain a pink color due to the thin layer of peptidoglycan. If a bacteria stains purple, due to the thick layer of peptidoglycan, the bacteria is a gram-positive bacteria. [4] In clinical microbiology numerous other staining techniques for particular organisms are used (acid fast bacterial stain for mycobacteria).