1. Shannon Proctor Departments of Chemistry and Biology Jacksonville University

2. Macrophages Greek for “big eaters” Originate from white blood cells called monocytes They play many roles in the body Major role in immunity and the immune response

3. Electron micrograph of a macrophage attacking bacteria. Representation of macrophages that I worked with using an inverted microscope. www.hartnell.edu/biology

4. Background Garlic has been used since ancient times as an herbal remedy. Heart disease Tumors The Plague Antibacterial properties

5. Background Previous research has shown that garlic enhances macrophage activity. Leishmania major, Ghazanfari 2005 Augmentation of function, Lau 1991 Enhanced immunocompetence, Lamm 2001 Our lab has shown that garlic does increase phagocytosis in macrophages

6. Internalization vs. Digestion Macrophages will phagocytose many things Engulf the particle May or may not destroy There is a chemical change when they digest O 2 uptake increases Rapid release of reactive oxygen species This is what we want to measure

7. Abstract An increase in the respiratory burst would be significant Nitro-blue tetrazolium (NBT) Yellow solution that turns blue

8. Cell Culture IC-21 Cells Cultured in RPMI media Incubated at 37°C, 5% CO 2

9. Phagocytosis Assay Subcultured at 100% confluency Detached Resuspended in control and media with garlic (0.5% or 1%) Incubation for 3hrs Addition of latex beads and fresh media

10. Phagocytosis Assay Repeated every 30, 60, and 90 minutes Incubation throughout Assay stopped by washing Staining using Hema 3 kit Inverted microscope for viewing 175-250 cells were counted and analyzed

11. 8-Well Slide

13. Results

14. Percent phagocytosis measured using the following equation: (# of macrophages with engulfed bead ÷ total # macrophages) X 100 D. Lindsey and K. Jackson, 2006 * = Control (60 % ethanol) = Garlic * P=.055, Student’s T-test

15. Results NBT Test Dissolve 1 tablet in water Fix cells (no staining) Cover cells with NBT solution and incubate at 37°C for 30 minutes Rinse with filtered PBS Observe color under microscope

16. Conclusion Future research could use latex beads to ensure phagocytosis and use a bacterium, for example, to measure the respiratory burst

17. Acknowledgements Dr. Karen Jackson, advisor Patricia Roman and Curtis Dobrowolski Jacksonville University Public Safety

18. References Buescher, E., Alling, D., and Gallin, J. (1985) Use of an x-linked human neutrophil marker to estimate timing of lyonization and size of the dividing stem cell pool. Journal of Clinical Investigation 76 (4), 1581-1584. Ghazanfari, T., Hassan, Z., and Khamesipour, A. (2005). Enhancement of peritoneal macrophage phagocytic activity against Leishmania major by garlic (Allium Sativum) treatment. Journal of Ethnopharmacology 103 (3), 333-337. Lindsey, D. and Jackson, K. (2006). Allium sativum Effects on Phagocytic Activity of IC-21 Peritoneal Macrophages in vitro