How to do a Gram Stain

1. Heat-fix a smear of the bacterial culture on a slide by passing the slide briefly through a small flame.
2. Apply crystal violet dye to the stain and wash off after 30 seconds. This is the primary stain.
3. Cover the rinsed slide with iodine, which is a mordans and will help crystalize the violet in the cells., after 30 sec, rinse off the iodine.
4. Wash the slide in alcohol to decolorize the slide.
5. Stain with a dark red solution of safranin which will serve as a counter stain.
6. Observe under a Lightfield Microscope, the bacteria which remained purple after the alcohol rinse is Gram-positive, the bacteria that is dark red (from the safarin) stain is Gram-negative.

Happy staining!