In Gram's procedure, the sample cells are first stained dark blue, followed by a thin cell wall decolorization step that is followed by a red dye counterstain.
- Crystal violet is kept inside of cells with a thick cell wall, giving the appearance of blue (gram positive) cells.
- In Gram negative bacteria, there is a thin peptidoglycan layer sandwiched between two thin cell membranes.
- The bacteria are first rinsed with crystal violet, a purple stain, then iodine to perform a Gram stain.
- Iodine and crystal violet combine to generate massive complexes that adhere to cells and color them purple.
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