Gram Stain Protocol

1. Make a smear of the bacterial broth on a clean glass microscope slide using aseptic technique.
2. allow to air dry
3. heat fix bacteria to slide by passing slide, bacteria side up, through flame 3 times
4. cover bacterial smear with crystal violet, sit for at least one minute
5. rinse briefly and blot
6. cover with gram iodine for at least one minute
7. rinse briefly
8. decolorize by dripping ethanol across surface of slide (7-15 drops or until first drop of E+OH is clear, not purple)
9. cover with safranin for one minute
10. rinse briefly and blot dry

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