User:Sydney Marshall/Notebook/Protease Project/2015/09/29

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Objective

  • Conduct Bradford Analysis between 1 μM Proteinase K and AuNP fibers

Description

The detailed protocol can be found in this notebook entry by Dr. Hartings.

  • AuNP Fibers Preparation
    • 7 eppendorf tubes with AuNP were centrifuged at 1500RPM for 1 min.
    • Their supernatant was aspirated
  • Protinase K 1 μM Preparation
    • Calculations
      • (0.00087g)(1mol/28900g)(1/0.001L)=30.1040 μM Proteinase K
      • M1V1 = M2V2 => (30.1040 μM)*(V1) = (1μM)*(1mL) => V1 = 0.0332mL = 33.2 μl Proteinase K (966.8 μl of buffer needed to add to make 1mL).
  • Incubation
    • 7 samples were incubated in a 37°C shaking water bath for 10 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hrs, 1.5 hrs, and 2 hrs each.
  • Bradford Analysis
    • At each time point, the samples were taken out of the water bath and centrifuged at 13500 RPM for 1 minute.
    • 750 μL of each sample was placed in a plastic cuvette with 600 μL of pre-mixed Bradford dilution and 1650 μL of buffer.
    • Samples were run from 400nm to 800nm on UV-Vis.


Notes

  • Blanks for this Proteinase K concentration were created and analyzed the next day.