Three New Nanotechnology Activities Recently Developed at the Minnesota Nano Center. James Marti, Ph.D. University of Minnesota Nano Center

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1 Three New Nanotechnology Activities Recently Developed at the Minnesota Nano Center James Marti, Ph.D. University of Minnesota Nano Center MicroNanoTechnology Conference June 24, 2015

2 The Minnesota Nano Center MNC) Nano-Link partner Interdisciplinary facility for research and prototyping Micro- & nanoelectronics, MEMS Nanomaterials Bio-nano systems

3 Research Experience for Teachers NSF Program Targets: HS and CC instructors Lab component Each teacher joins a research group Attend conferences, co-author papers,

4 Research Experience for Teachers Lesson development component Select a nanoscience topic Outline, first draft, second draft Field test to refine (fall) Final version posted to NNIN Education Portal

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6 MNC s RET Program Run Seven HS, two CC teachers Two repeat participants Total of ten lessons posted to web site Fully developed and classroom tested supply lists, sources student guide with questions connection to state science standards

7 Featured Nanoscience Lessons Brine Shrimp and Nanosilver (2012) Using Gold Nanoparticles for Bacterial Detection (2013) Spectrophotometry with Metal Nanoparticles (2014)

8 The Effects of Gold and Silver Nanoparticles on Brine Shrimp: A Toxicology Study Sharon Meierhofer 1, Melissa Maurer-Jones 2, Sara Love 2, Zhen Liu 2, Christy Haynes 2, and James Marti 3 1 Southwest High School, Minneapolis, MN 2 Department of Chemistry, University of Minnesota, Minneapolis, MN 3 Nanofabrication Center, University of Minnesota, Minneapolis, MN Published in Journal of Chemical Education, April 2013 (Vol.90(4), pp )

9 Background Silver nanoparticles are used specifically for their antimicrobial effects. This lab allows students to study the toxicity of gold and silver nanoparticles on an example organism, brine shrimp.

10 Lesson Overview Students synthesize and prepare several dilutions of gold and silver nanoparticles to determine toxicity on brine shrimp. Target grade levels: High school Traditional science classes: biology, chemistry, environmental science

11 Student Motivation Products using nanoparticles are used in household products sunscreen: zinc oxide clothing and food storage: silver What is the potential impact of these nanoparticles on organisms in the environment?

12 Goals of lab This lab is designed to introduce students to a practical application of chemistry in the real world the field of nanotoxicology as an interdisciplinary field of science the need to carefully conduct experiments and evaluate data in a collaborative environment to measure toxic hazards.

13 Materials and Equipment High school chemistry lab equipment brine shrimp eggs and growth medium aquarium air pump gold nanoparticle synthesis reagents silver nanoparticle synthesis reagents 24-well plates for shrimp viability testing bovine serum solution

14 Procedure Instructor Advance prep hatch brine shrimp (2-3 days ahead) prepare gold and silver nanoparticle stock solutions Make serum solution

15 Visual Summary of Procedure

16 Work Plan Day Day 1: Nanoparticle synthesis and dilution Tasks 1) Carry out Au and Ag NP syntheses (UW MRSEC procedure) 2) Perform serial dilutions of NP dispersions Day 2: Brine shrimp counting and exposure 1) Conduct live shrimp assay 2) Expose shrimp to assigned NP concentrations Day 3: Counting surviving brine shrimp and data analysis 1) Conduct live shrimp assay. 2) Perform statistical measures on their own data 3) Combine data into group data and then classroom data. 4) Complete graphs and questions as homework

17 TEM images of Au and Ag nanoparticles prepared by students in this experiment

18 Image of brine shrimp eggs (inset) and live brine shrimp

19 Nanoparticle Exposure Tests

20 Nanoparticle Exposure Tests Place different concentrations of Au and Ag nanoparticles in a 24 well plate Include solution with dissolved silver (ions, not NPs) Add brine shrimp Manual census: how many alive (moving)? Incubate for 24 hrs Recount shrimp

21 Viability Calculation viability ( in %) = # live brine shrimp after exposure x 100 #live brine shrimp before exposure

22 Viability Data Nanoparticle Concentration Initial number of brine shrimp (before exposure to nanoparticles) Number of live brine shrimp (after exposure to nanoparticles) % Viability of brine shrimp control Ion Control 100 % Gold nanoparticles 50 % Gold nanoparticles 25 % Gold nanoparticles 12.5 % Gold Nanoparticles

23 Expected Results Would expect the brine shrimp to show least viability in the 100% silver nanoparticles increasing viability in the 50%, 25% and 12.5 % silver nanoparticles The gold nanoparticles are not expected to impact viability.

24 Classroom Testing Three classes of high school chemistry students performed this experiment (2 standard, 1 AP) Each were given a five question test prior to and following the experiment Tested comprehension of Nanoscience concepts (scale, etc.) Nanoparticles in products How to test for hazardous substances Statistical reasoning The interdisciplniary nature of toxicology

25 Testing Results 1. Based on pre- and post-testing answers, the students showed improvement in understanding of nanoscience concepts, nanoparticles, and toxicology as an interdisciplinary science limited improvement in using statistical measures (responses largely unchanged after performing the experiment)

26 Testing Results 2. Students had a difficult time counting the live brine shrimp count difficulty in the fast movement of the shrimp Protoslo" ( a matrix to slow down the shrimp) was used, but with no better results. The effect of the inaccurate pre- and post-live count was that statistical analysis didn't clearly show expected results.

27 For Future Work For more accurate brine shrimp counts, each group of students could disperse individual brine shrimp in separate well plates and treat each brine shrimp with exposure to test concentrations separately. Sufficient sample size will be obtained due to the large number of groups collecting data. Data would again be shared among all classes for analysis. Discussion: Should the use and disposal of silver nanoparticles be regulated?

28 Using Gold Nanoparticles for Bacterial Detection Mike Falck 1 and James Marti 2 1 Chaska High School, Chaska MN 2 Minnesota Nano Center, University of Minnesota, Minneapolis, MN

29 Gold Nanoparticles in Medicine Can target a wide variety of biological materials, Can be used for bio assays and other diagnostic tests delivery of specific drugs and the destruction of tumors. Surface modified GNPs expensive, complicated to work with

30 Using Gold Nanoparticles to Simulate Bacterial Detection Students create gold nanoparticles (GNP s) approximately 15 nm in size. These GNP s will be used to detect simulated meningitis bacteria which will infect some of your students! Part II of this lab will consist of all students kissing other students (bacterial meningitis is only contagious through bodily fluids)! Students simulate swapping spit by using test tubes and eye droppers. Students then use their engineered GNP s to detect those who have contracted the disease.

31 Activity Overview In this lab students will become familiar with gold nanoparticles As part of the lesson students will learn the potential disease detection uses of GNP s, conduct research on modifying GNP s to be specific to other molecules, and complete a simulation using GNP assay tests to detect a simulated meningitis (Neisseria meningitides) outbreak in the class. Target grade levels: Traditional science classes: May be used in Biology or Biomedical classes

32 Materials and Equipment Part I- Making Gold nano particles HAuCl 4 - Chloroauric acid Na 3 C 6 H 5 O 7 -Sodium citrate NaCl-table salt 50 ml Erlenmeyer Flasks 100ml Erlenmeyer Flasks Test tubes Eye droppers (graduated if possible) Hot plate/stirrers Part II- GNP Assay Test Simulation. 1 test tube with NaCl solution (simulated antigen, given to one student) Everybody else receives test tubes with distilled water only Each student needs an eye dropper GNP product from part 1

33 Procedural Breakdown Time Activity Goal Day 1 The day before the lab 48 min Introduce students to the topic of nanoparticles. Discuss gold nanoparticles specifically. Have students research uses, and share with class. To prepare students to create and use gold nanoparticles for medical uses as an assay test. This lab is intended to connect GNP s to the medical field. The lab simulates a meningitis assay test. Feel free to change the disease to whatever fits your curriculum. Review how assay tests are used. Prepare lab material for part I and II.

34 Procedural Breakdown Time Activity Goal Day The day od student lab part I 5 min Students answer warm-up questions. what are gold nanoparticles? To ensure students understand what gold nanoparticles are and how they can be used specifically as assay tests. discuss three uses for gold nanoparticles with your partners min Distribute Student Worksheets to students. Students follow procedures and complete Part I of the lab. To allow students to complete part I of the lab. Any extra time could be spent reviewing assay tests. 5min Clean up. To prepare workspace for next class.

35 Procedural Breakdown Day 3 Time Activity Goal The day of student lab part II 5 min Students answer warm-up questions. what are gold nanoparticles? To ensure students understand what gold nanoparticles are and how they can be used specifically as assay tests. Why is part II of this lab only a simulation 35-40min What should we notice in our samples when the GNP is added? Instruct student to take out Student Worksheets. Students follow procedures and complete Part II of the lab To allow students to apply their GNP s to solve a problem. Who has meningitis and who started the outbreak? 5min Clean up. To prepare workspace for next class.

36 Procedural Breakdown Day 4 (optional) The day of the student lab part III 5 min Students answer warm-up questions. what are gold nanoparticles? To ensure students understand what gold nanoparticles are and how they can be used specifically as assay tests. How did GNP allow us to detect infected people yesterday? 35-40min How was yesterday s lab realistic and unrealistic? Instruct student to take out Student Worksheets. Students follow procedures and complete Part III of the lab To allow students to apply their GNP s to solve a problem. How can GNP s be modified to be used for nanoscience? 5min Clean up. To prepare workspace for next class. Day 5 The day of the student lab part III (optional) 45 min Students present findings from previous day s research.

37 Teacher Pre-lab Prep Procedures Part I- Producing GNP s. 1. Prepare 500 ml of 1.0 mm HAuCl4 solution by dissolving 0.1 g of HAuCl4 in 500 ml of distilled water. This solution is unstable and keeps only a few days, so make it fresh for the lab. 2. Prepare 50 ml of 38.8 mm Na 3 C 6 H 5 O 7 (Sodium Citrate)solution. Part II- Assay Test Simulation 1. Prepare a 1 M solution of NaCl by dissolving 0.5 g of NaCl in 10 ml of distilled water to be used as antigen. (This will act as a meningitis vector for the simulation)

38 Student Procedure Part I (Summarized) Part I- Making GNP s Students obtain 20ml 1.0 mm HAuCl4 solution, place it in an Erlenmeyer flask on a hot place, heat to boiling and add add 2 ml of citrate solution. After boiling and stirring the solution, a deep red color appears. This takes about 10 minutes.

39 Student Procedure Part II (Summarized) Part II Student receive an eye dropper and at a test tube filled with 10 ml of distilled water EXCEPT for one student who needs to receive the meningitis sample (NaCl solution) Students kiss two other people (as meningitis is contagious via body fluids). To kiss students will squirt 2ml of their test tube solution into a partner s test tube and vice versa.

40 Student Procedure Part II (Summarized) Students will add 1.5 ml of their GNP from day 1 to perfom assay test. Optional-Students can play the part of an epidemiologist by figuring out who was the original vector of the disease.

41 Student Procedure Part III (Summarized) Part III (optional) Students research real world methods of treating GNP s for use in the medical field and present there findings

42 Analysis A color change will signal a positive result for meningitis (positive results should change blue). No color change or a slight brightening of the pink red signals a negative result for meningitis. Students can conduct an investigation to determine who carried original pathogen.

43 Going Further Students research one way in which GNP surfaces can be modified to be specific to a molecule (ex specific strain of DNA, RNA or protein) and present findings.

44 Results of Field Testing: Students from the Chaska High School Medical Interventions class performed this experiment, and were given conclusion questions, took part in class discussions and took a 15 question quiz following the experiment to assess their comprehension of the core themes of the activity.

45 Activity Update The classroom trial helped to refine the final version of the experiment changes were made to the division of experiments and to the gold nanoparticle synthesis.

46 Spectrophotometry to Estimate the Size of Student-Made Nanoparticles Chris Kaus 1, Damini Gupta 2, Kevin Dorfman 2, and James Marti 3 1 Richfield High School, Richfield, MN 2 Department of Chemical Engineering and Material Sciences, University of Minnesota, Minneapolis, MN 3 Minnesota Nano Center, University of Minnesota, Minneapolis, MN

47 Lesson Overview Students create either gold or silver nanoparticles in class. Students use a spectrophotometer to create % transmittance and absorbance curves of their samples. Students compare their curves with three absorbance curves from commercial-grade standards to estimate the size of their nanoparticles. After reading an article from ChemMatters on nanoparticles, students then research other uses and applications of either gold or silver nanoparticles. Target grade levels: High school chemistry

48 Goals of lab Introduce students to nanoparticle properties and technology the use of a spectrophotometer for quantitative analysis comparison of lab results using interpolation and/or extrapolation to estimate a quantitative value.

49 Materials and Equipment Gold and silver nanoparticles were fabricated using the University of Wisconsin MRSEC procedures. For production of silver nanoparticles: magnetic stirrer, stir bar, bowl for ice bath, small Erlenmeyer flask, 50 ml graduated cylinder, dropper, vial for storing nanoparticle solution 30 ml of 0.002M sodium borohydride (NaBH 4 ) 2 ml of 0.001M silver nitrate (AgNO 3 ) For production of gold nanoparticles: Stirring hot plate, stir bar, ml Erlenmeyer flask, 25 ml graduated cylinder, 10 ml graduated cylinder, container for storing nanoparticle solution 20 ml of 1.0M HAuCl 4 2 ml of a 1% solution of trisodium citrate dihydrate, Na 3 C 6 H 5 O 7 2H 2 O

50 Materials and Equipment Spectrophotometer to create % transmittance and absorbance curves Laser to test for the presence of a colloidal suspension

51 Procedures Students create either silver or gold nanoparticles Silver NP being stirred in an ice bath Students waiting for their gold NP to boil.

52 Spectrophotometry of Silver/Gold NP Students using spectrophotometers to create % transmittance and absorbance curves Students using the analog spectrophotometer from the 1960 s! Students using the new digital spectrophotometer

53 Transmittance and Absorbance Curves Curves obtained by a group of students who produced silver nanoparticles.

54 Transmittance and Absorbance Curves Curves obtained by a group of students who produced gold nanoparticles.

55 Absorbance Curves of Standards (provided by NanoComposix, Inc.) 10 nm Gold Nanoparticle Standard 60 nm Gold Nanoparticle Standard 100 nm God Nanoparticle Standard 10 nm Silver Nanoparticle Standard 60 nm Silver Nanoparticle Standard 100 nm Silver Nanoparticle Standard

56 Expected Results The class expected very little variation in the sizes of silver NP and gold NP. All silver transmittance and absorbance curves will be similar, as they will with the gold NP.

57 Analysis of Results Students who created the silver nanoparticles had similar results. Students estimated the size of their silver nanoparticles to be between nm. Students who made gold nanoparticles were not able to create transmittance or absorbance curves. It appeared the gold NP solution was too dark for the spectrophotometers. Sample gold NP solutions were given to the students to analyze to create absorbance curves. Results of this analysis showed a consistent size of approximately 70 nm.

58 Analysis of Gold NPs U of M used a dynamic light scattering tool to analyze our gold NP sample

59 Surprises from Activity Most of the activity went very smoothly. We were not able to analyze the gold nanoparticles in our spectrophotometers. Not exactly sure the reason. The solution was a very dark red. Perhaps too dense.

60 Modifications for Next Year Have students create their own absorption graphs from the standard solutions. Provides practice in using a spectrophotometer Allows students to compare their results directly with others (results should be similar) Students should maintain detailed records of their synthesis process. For example, record exactly how long they stirred the silver NP in the ice bath and the time needed for the gold NP to turn a deep red. This will lead to discussion about varying results between groups. May assist in resolving why we were not able to analyze the gold NP.

61 Modifications for Next Year There are several great Youtube videos showing the applications of nanoparticles. Nanoparticle drug delivery in cancer therapy 7 surprising facts about silver nanoparticles and health Gold nanoparticles: strange properties and applications

62 For more info The Minnesota Nano Center: mnc.umn.edu Characterization Facility: charfac.umn.edu James Marti Minnesota Nano Center Phone: (612)

63 A. Who we are and what we do B. Technical capabilities C. Recent work at the MNC D. Ways to work together E. Other R&D resources at the University