NimitMistry13
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Having read the gram-stain process, I don't understand why the bacteria are not killed by the alcohol wash.
From my understanding:
1) the sample is stained with crystal violet
2) iodine is applied to fix the dye
3) stain is washed with alcohol (washing stain off gram-negative bacteria)
4) sample is stained with safranin
5) Only gram-negative bacteria, which were colourless before, become pink from safranin

How does the alcohol wash off the crystal violet stain, but not kill the bacteria if it dissolves their cell wall? Is it a low concentration or something?
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HarisMalik98
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Gram staining is dependant on the cell wall composition, not the viability of the bacteria. Most of the bacteria will be killed during the heat-fixation stage at the very start; and almost all will be killed by the end of the staining procedure. However, to work, the process itself only requires the presence of the cell wall - so, whether the bacteria are alive or not during the procedure is irrelvant.

So to answer your question; most bacteria will already be dead before the decolourisation stage, however, the alcohol may kill bacteria that survived previous stages. But as I said, it doesn't matter.

Hope this helps.
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NimitMistry13
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(Original post by HarisMalik98)
Gram staining is dependant on the cell wall composition, not the viability of the bacteria. Most of the bacteria will be killed during the heat-fixation stage at the very start; and almost all will be killed by the end of the staining procedure. However, to work, the process itself only requires the presence of the cell wall - so, whether the bacteria are alive or not during the procedure is irrelvant.

So to answer your question; most bacteria will already be dead before the decolourisation stage, however, the alcohol may kill bacteria that survived previous stages. But as I said, it doesn't matter.

Hope this helps.
Thank youuuuu
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