1. Cytotoxic and Immunologic Effects of Silver Nanoparticles Shaun Cote

2. What are Nanoparticles?  Any substance that has one dimension of 1-100 nanometers  Commonly used nanoparticles include colloidal gold and titanium dioxide (TiO 2 )

3. Nano Scale

4. Why is this important?  “The biokinetics of NSPs are different from larger particles. When inhaled, they are efficiently deposited in all regions of the respiratory tract; they evade specific defense mechanisms; and they can translocate out of the respiratory tract via different pathways and mechanisms (endocytosis and transcytosis).” Nanotoxicology: An Emerging Discipline Evolving from Studies of Ultrafine Particles. Environmental Health Perspectives, Oberdörster et al. 113(7), July 2005.

5. Downsides to Silver Nanoparticles  Possible negative effects due to the chemistry of small molecules

6. How they are Made  Metal salt solutions are boiled and then reduced via a reducing agent

7. Silver Nanoparticles in the Market  Samsung  Washing machine, refrigerator and air conditioners  Nanohorizons  Smart Silver – silver nanoparticles intertwined with fabric to repel odor and microbes  Aigon  SilverClene anti-microbial cleaning solution

8. Project Goals  To explore the effects of silver nanoparticles by testing the:  Cytotoxicity on established cell lines:  Production of specific immune molecules

9. Testing 1) MTT Assay – tests for metabolic activity 2) TNF-α ELISA – Antibody test for TNF-α secretion from cells 3) RT-PCR – Measure of RNA activity

10. Cell Lines Tested  RAW 264.7 – Mouse macrophage  Jurkat – Human T cells  WHTBF-6 – Human lung fibroblast

11. MTT Assay  Cells are cultured, then treated with silver nanoparticles  MTT is added, and blue crystals are formed if the cells are viable  Correlation between concentration of nanoparticles and viability of the cells

12. MTT Assay - Results

13. TNF-α ELISA  Test based on antibody capture of TNF-α, a protein molecule essential in triggering an immune response  Measured OD of sample wells compared to a standard curve

14. TNF-α ELISA Results  Concentration of silver nanoparticles versus relative concentration of TNF-α

15. RT-PCR  A method of amplifying target RNA to determine how silver nanoparticles affect gene expression of specific cytokines

16. Results of RT-PCR  Only tested GAPDH, a housekeeping gene  Further tests include the cytokines TNF-α, IL-6, and IL-1β

17. Further Studies  Continuing RT-PCR studies on other cytokines

18. Acknowledgements  Dr. Ah-Kau Ng and Lab  Theresa Knight  Jennifer Walker  SURF program