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The decolorization step is critical and must be timed correctly; the crystal violet stain is removed from both gram-positive and negative cells if the decolorizing agent is left on too long (a matter of seconds). [19] After decolorization, the gram-positive cell remains purple and the gram-negative cell loses its purple color. [19]
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. The Gram stain is used by microbiologists to place bacteria into two main categories, Gram-positive (+) and Gram ...
Unlike negative staining, positive staining uses basic dyes to color the specimen against a bright background. While chromophore is used for both negative and positive staining alike, the type of chromophore used in this technique is a positively charged ion instead of a negative one. The negatively charged cell wall of many microorganisms ...
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.
There is no connection between the shape of a bacterium and its color upon Gram staining; there are both gram-positive rods and gram-negative rods. MacConkey agar can be used to distinguish among gram-negative bacilli such as E. coli and salmonella. [23]
One commonly recognizable use of differential staining is the Gram stain. Gram staining uses two dyes: Crystal violet and Fuchsin or Safranin (the counterstain) to differentiate between Gram-positive bacteria (large Peptidoglycan layer on outer surface of cell) and Gram-negative bacteria. Acid-fast stains are also differential stains.
Examples include the malachite green counterstain to the fuchsine stain in the Gimenez staining technique and the eosin counterstain to haematoxylin in the H&E stain. [1] In Gram staining, crystal violet stains only Gram-positive bacteria, and safranin counterstain is applied which stains all cells, allowing the identification of Gram-negative ...
[2] [3] Carbol fuchsin is used as the primary stain dye to detect acid-fast bacteria because it is more soluble in the cells' wall lipids than in the acid alcohol. If the bacteria is acid-fast the bacteria will retain the initial red color of the dye because they are able to resist the destaining by acid alcohol (0.4–1% HCl in 70% EtOH). [4]