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

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

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

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

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

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

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

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

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

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

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

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

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

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

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