How Does Culture Age Affect the Results of a Gram Stain?


Quick Answer

Culture specimens that are over 24 hours old may stain incorrectly, losing their ability to hold on to the crystal violet stain. They may appear as gram-negative organisms when they are actually gram-positive, or the specimen may show a gram-variable result.

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Gram stains reveal the structural differences in the cell walls of organisms in the laboratory. Gram-positive organisms have a thick cell wall containing up to 90 percent peptidoglycan and appear blue or purple under the microscope after staining with crystal violet. Gram-negative organisms have a much thinner cell wall containing up to 10 percent peptidoglycan and a high lipid content. These organisms appear red or pink under the microscope after staining with crystal violet.

The gram stain is a critical laboratory test that is integral to the study and treatment of infectious disease. Many antibiotics work on either gram-negative or gram-positive organisms because of the way they break down the cell walls. Broad-spectrum antibiotics work on both gram-negative and gram-positive organisms.

The gram stain was created by Hans Christian Gram in 1884. He was searching for a way to visualize cocci in lung tissue samples of those who had died from pneumonia. He used crystal violet as his primary stain, followed by iodine as a fixer and ethanol as a decolorizer. The decolorizing step creates the loss of the blue-purple coloration that differentiates gram-positive and negative organisms from one another.

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