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Cell wall has different organization in Gram-positive and Gram-negative bacteria. In Gram-positive bacteria the cell wall is represented mainly by peptidoglycan molecule, which is arranged in a number of layers and creates a strong carcass for the cell. The thickness of cell wall in Gram-positive bacteria varies from 20 to 80 nm. In Gram-negative bacteria the layer of peptidoglycan is much thinner. However, it is not the only component of cell wall. A thick layer of lipoproteins is present over the layer of petidoglycan. Difference in the organization of cell wall in Gram-positive and Gram-negative bacteria results in the difference in staining. Difference between the two groups observed during Gram staining is based on the ability of stains belonging to the group of triphenylmethane to form complex with peptidoglycan in the presence of iodine. Gram-positive bacteria are stained purple or dark blue because triphenylmethane is not washed with alcohol. But in case of Gram-negative bacteria the complex is washed with the application of alcohol. Thus, they are stained with saphranine applied after alcohol and appear red.
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