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Development of Rapid Through-put Nanotoxin Bioassays

Development of Rapid Through-put Nanotoxin Bioassays. Perry Kanury Cassandra Viéville ~ Dr. Stacey Harper CBEE/EMT. Background:. Nanomaterials are widely used in both industrial and consumer applications. Currently thousands of distinct compounds in use, with more introduced every day

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Development of Rapid Through-put Nanotoxin Bioassays

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  1. Development of Rapid Through-put Nanotoxin Bioassays Perry Kanury Cassandra Viéville ~ Dr. Stacey Harper CBEE/EMT

  2. Background: • Nanomaterials are widely used in both industrial and consumer applications • Currently thousands of distinct compounds in use, with more introduced every day • Found in every day products such as deodorant, sunscreen, Teflon, pesticides, etc. • Little or no knowledge of the true environmental or molecular impacts of these materials • Distinct lack of methods or protocols for toxicity testing

  3. What are nanomaterials? • Materials with at least one dimension between • 1 and 100 nanometers • Planar: Single dimension in ‘nano’ scale • Example: Graphite sheet • Tubular: Two dimensions in ‘nano’ scale • Example: Bucky Tube • Globular: Three dimensions in ‘nano’ scale • Example: Bucky Ball

  4. What are nanoparticles? • Subcategory of nanomaterials • Formerly known as 'Ultra Fine Particles' • At least two dimensions between 1 and 100 • nanometers in diameter, implying either tubular or • globular shapes • Can take forms such as 'Nanoclusters', 'Nanocrystals', or 'Nanopowders' depending on agglomeration behavior • Wide range of applications including biomedical, optical and electronic fields • Often exhibit unique structure-property characteristics • as a result of volume to surface area ratio and • surface structure • For reference: A typical nanoparticle has an effective diameter of approximately 1/800th that of a human hair.

  5. What does this mean to us? • Wide range of possible effects to both human and ecological systems Examples: • Introduction of antimicrobial agents such as Aluminum Zirconium Tetrachlorohydrex (active ingredient in deodorant) into a town’s microbe-based water treatment system and subsequent watershed • Potential endocrine disruption (among additional myriad of effects) upon chronic exposure to Titanium Dioxide (active ingredient in many sunscreens) near lymph nodes, etc. • Possible disruption of ‘Lock and Key’ hormone identification mechanisms within the Endocrine system.

  6. Need: • Rapid testing strategies are necessary to identify specific nanomaterials that result in toxicity in order to mitigate risks from exposure and define structure-property relationships that can be used to predict nanomaterial fate and hazard in lieu of empirical data.

  7. Hypothesis:  An aquatic ecosystem can be modeled in the form of a ‘nanocosm’ ~ an extremely small tritrophic ecosystem (250 uL). Prediction: This ecosystem can be effectively used for the purpose of assessing the toxicity of nanoparticles in aquatic environments, as well as modeling the impacts of these nanoparticles on said ecosystems.

  8. Another way of looking at it:

  9. Another way of looking at it: ‘Nano’ Algae    Ciliate    Bacteria (All 3 organisms) Varying nanoparticle exposures

  10. Tri-trophic aquatic ecosystem based upon our local temperate region • Photo-synthesizer (primary producer): Chlamydomonas reinhardtii (green algae) • Predator (primary consumer): Tetrahymena thermophilia (ciliate) • Decomposer (detritovore): E.coli (bacteria) • Suspended in a defined microbial media • Upon exposure by nanotoxins, can monitor population dynamics and predator/prey interactions to understand effects. The makeup and utilization of a Nanocosm: ‘Nano’ AlgaeCiliate    Bacteria (All 3 organisms) Varying nanoparticle exposures

  11. Typical Plate Layout Nano Bacteria Algae Ciliate Exposure Concentration 0.01 ppm 0.05 ppm 0.1 ppm 0.5 ppm 1 ppm 5 ppm 10 ppm 50 ppm Three replicates of each treatment for each organism

  12. How we observe these interactions: ‘Nano’ Algae   Ciliate    Bacteria (All 3 organisms) Flow Cytometer Quantifies and categorizes cells via light diffraction and  fluorescence.  Varying nanoparticle exposures Cell Concentration (cells/uL) of each organism are obtained and analyzed based upon effective size and fluorescence.

  13. How we observe these interactions: Tetrahymena Thermophilia AgNO3 Exposure over 5 days

  14. How we observe these interactions: Concentration Response Treatment is observed relative to negative control within the same timepoint (((treatment-control)/control)*100)

  15. Future Work: • Investigation of more ideal organisms than Tetrahymena thermophilia in order to streamline data collection utilizing existing equipment • Investigation of additional endpoints beyond mortality • Investigation of possible mechanisms of toxicity • Establishment of a database of the effects of various nanotoxin exposures.

  16. Acknowledgements: • Dr. Stacey Harper • Cassandra Viéville • Howard Hughes Medical Institute • Environmental Health Sciences Center (EHSC) • Oregon State University • Dr. Kevin Ahern

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