Gram Stain Essay

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Exercise 10 Gram Stain Gram staining, a differential staining technique, is one of the most important methods used by microbiologist. The technique allows us to separate bacteria into two fundamental groups; Gram positive and Gram negative. The differences between these two groups rests in the chemistry of their cell wall. Gram positive cells have a cell wall composed of a very thick layer of peptidoglycan, a uniquely bacterial molecule. While, Gram negative bacteria have a cell wall composed of a thin layer of peptidoglycan surrounded by an outer membrane. Hans Christian Gram, developed the method in 1884 while trying to detect bacterial cells in mammalian tissue. By exposing the tissues to crystal violet (a basic stain) and then rinsing with a solvent, Gram, found the bacteria retained the purple color of the crystal violet while the tissues did not. He then flooded the tissue with safranin (a basic stain) as a counterstain to dye the cells that had been decolorized by the solvent. Safranin, a mild red color, is of much lower intensity then crystal violet, therefore, it does not change the color of the purple stained cells but it will stain those that are clear a light red. Gram soon learned that not all bacterial cells retained the crystal violet after the solvent rinse. Some bacterial cells would be rinsed clear only to be stained red by the counterstain. These bacteria are called Gram negative. When bacteria are flooded with crystal violet the stain binds to cell wall molecules of all bacteria, like any simple stain will, staining them all purple. Because Gram positive bacteria have a thick peptidoglycan layer (30 layers or more) the stain penetrates deep into the meshwork of these molecules. Gram negative bacteria only have one or a few layers of peptidoglycan molecules. When iodine is added it acts as a mordant (fixes dye), complexing with the
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