GRAM STAIN PROCEDURE Ø 1 Gather all equipment

GRAM STAIN PROCEDURE Ø 1. Gather all equipment and supplies – Gram stain – Microscope slides – Heat source – Sterile swab – Culture or specimen – Microscope

GRAM STAIN PROCEDURE Ø 2. Label the slide with the patient’s initials or the specimen number.

GRAM STAIN PROCEDURE Ø 3. Choose an isolated colony off of the agar plate and obtain bacteria with a sterile swab.

GRAM STAIN PROCEDURE Ø 4. Place the swab on the microscope slide and spread the colonies in a circular motion.

GRAM STAIN PROCEDURE Ø 5. Heat fix the microorganisms to the slide by placing the bottom of the slide to heat for approximately 30 seconds.

GRAM STAIN PROCEDURE Ø 6. Place slide on staining tray or hold with forceps above the sink.

GRAM STAIN PROCEDURE Ø 7. Flood the surface of the slide with Crystal Violet stain and let sit for one minute.

GRAM STAIN PROCEDURE Ø 8. Rinse the slide with distilled water.

GRAM STAIN PROCEDURE Ø 9. Flood the slide with Gram’s Iodine and time for one minute.

GRAM STAIN PROCEDURE Ø 10. Rinse the slide with distilled water.

GRAM STAIN PROCEDURE Ø 11. Flood the slide with Gram’s decolorizer and time for 30 seconds.

GRAM STAIN PROCEDURE Ø 12. Rinse the slide with distilled water.

GRAM STAIN PROCEDURE Ø 13. Flood the slide with the counterstain, Safranin, and let sit for one minute.

GRAM STAIN PROCEDURE Ø 14. Rinse the slide with distilled water.

GRAM STAIN PROCEDURE Ø 15. Blot the slide and read with the oil immersion lens of the microscope. Look for Gram-negative and Gram-positive bacteria.