IGEM:American University/2009/Notebook/BCHM 2/2017/03/01

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Sample Preparation

Solutions for lysozyme templated quantum dots were prepared in a 5 mL rxn mixture as follows:

Reagents

  1. 0.5 M Potassium thioacetate
  2. 200 uM Lysozyme in Water
  3. 5 mM Cadmium acetate in Water
  4. Ammonia water for pH adjustment
  5. HNO3 for pH adjustment
  6. Deionized Water

Volume Additions

  1. 100 uL Potassium Thioacetate
  2. 312.5 uL Lysozyme
  3. 1000 uL Cadmium Acetate
  4. 3587.5 uL DI Water
  5. <10 uL HNO3 or Ammonia water for pH adjustment

It should be noted that we had issues with sulfur precipitating in the plastic scintillation vials. This may have been because of the plastic interacting with the sulfur. I also believe that it may have something to do with the order of the sulfur source addition. In the article, “Cadmium Sulfide Nanoparticle Synthesis in Dps Protein from Listeria innocua,” Kenji Iwahori discusses the reaction mechanism and requirements for the formation of CdS QDs within a protein template. It is not discussed in the Naito Article, "Circularly Polarized Luminescent CdS Quantum Dots Prepared in a Protein Nanocage," that the addition step should be the final step. The Iwahori article describes issues with sulfur crashing out when it is not added slowly, dropwise, and as the final addition step. This procedure added the potassium thioacetate as the final reagent; 100 uL of this reagent was added dropwise over the course of a 60s while the rxn mixture was stirring at 260 rpm on a stir plate. We believe that this will aid in proper formation of the nanoparticles and prevent sulfur crashing out. Sulfur suspended in the solution has in the past formed a colloid, which may lead to issues with the inner filter effect during fluorescence analysis. It is our hope that changing the order of the procedure will prevent this issue.

It should also be noted that because the potassium thioacetate is added as the final reagent, that the pH may be slightly affected. The pH study is done to show a range of pH, so the effect of the potassium thioacetate being added as the final reagent should change the pH of the rxn mixtures the same amount. This will result in some pH error that we will disclose in the final paper. This is not a major issue because the pH is meant to show range, not exact pH.


Link Iwahori: http://pubs.acs.org/doi/abs/10.1021/cm0628799 Link Naito: http://onlinelibrary.wiley.com/doi/10.1002/anie.201002552/abstract

Below is an image of the rxn solns after preparation. Day 0. It should be noted that the high pH solutions are cloudy. We will more thoroughly explore the cause of this next week. It may be because of the protein's AA interactions at high pH. It may also be because the sulfur source is affected by high pH. 3 1 17 CdS Lysozyme Day 0.jpg

Data Analysis Overview: ZnS QDs Templated via BSA

All together AnnelieseAnalysis030117.PNG All Together View AnnelieseAnalysis030117.PNG

Data Analysis In-Depth: ZnS QDs Templated via BSA

pH 4

PH 4 AnnelieseAnalysis030117.PNG

pH 4 Selected View

PH 4 ViewAnneliysis030117neliysis030117.PNG

pH 5

PH 5 AnnelieseAnalysis030117.PNG

pH 5 Selected View

PH 5 View AnnelieseAnalysis030117.PNG

pH 6

PH 6Anneliesefaustinoanalysis030117.PNG

pH 6 Selected View

PH 6 ViewAnneliesefaustinoanalysis030117.PNG

pH 7

PH 7Anneliesefaustinoanalysis030117.PNG

pH 7 Selevted View

PH 7 ViewAnneliesefaustinoanalysis030117.PNG

pH 8

PH 8 Anneliesefaustinoanalysis030117.PNG

pH 8 Selected View

PH 8 viewAnneliesefaustinoanalysis030117.PNG

pH 9

PH 9Anneliesefaustinoanalysis030117.PNG

pH 9 Selected View

PH 9 View Anneliesefaustinoanalysis030117.PNG

pH 10

PH 10Anneliesefaustinoanalysis030117.PNG

pH 10 Selected View

PH 10 ViewAnneliesefaustinoanalysis030117.PNG