February 11th
Goals
- We have to redo the dopamine addition that we did last Wednesday
- We are going to test the activity of citrate-HRP-AuNPs using O2 binding assays
Procedures
Activity of citrate-HRP-AuNPs
- We are going to use an assay provided by Dr. Hartings
- The assay uses a reagent that changes color when HRP is added in addition to hydrogen peroxide
- The hydrogen peroxide alone will change the color of the solution, so we will be comparing hydrogen peroxide alone with hydrogen peroxide and AuNPs
- We also are going to run UV Vis of the samples to see how the absorption spectra changes over time
Dopamine Coating
- Dissolve dopamine in 100mL Tris Buffer to have a 10mM solution at pH=8.5
- Mix 17mL of AuNP solution with 17mL of the dopamine solution
- Stir under air at room temperature for 1 hour
- Purify by centrifuging and then redisperse in water.
- Filtrate through syringe filters with cellulose acetate membranes with pore size of 1um
- Make 2 solutions.
- to be stored over night on lab bench
- to be stored over night in the refrigerator
Notes
- the nanoparticles with HRP were functional
- In the 10^-11 dillution, we observed a color change from red initially to a much lighter solution
- In our calculations we decided to use teh 10^-11 and water as our blank, so we could subtract protein + water.
Protein, OBD, H202
- For each sample run in UV-Vis there was 0.225ml HRP, 2.25ml OBD stock and 0.75ml hydrogen peroxide stock
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