User:Monika Gasiorek/Notebook/CHEM-571 2014F/2015/04/01

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April 1st, 2015

1 μM Proteinase K Reaction w/ New Protocol

  • 5 - 5 mL test tubes of AuNP fibers were vortexed to produce "homogenous" fiber solutions.
  • 20 - 1.5 mL epitubes were each filled with .927 mL of the fiber solution and 0.020 mL of a concentrated 2.5 M Tris-HCl/0.5 M CaCl2 pH 8 buffer.
  • For the (approx) first half of the samples being obtained (0.5, 1, 1.5, 2, 3, 4, 5, 6, 7, 8, 9 min), 0.053 mL of the 19.03 μM proteinase K stock solution was added to each epitube at separate points in time, letting each sample run to its specific timepoint, at which point the epitube was centrifuged for 30 seconds and a 500 μL sample was extracted before adding the protease to another fiber sample to let it run for its respective time.
  • The 0.053 mL of proteinase K was added to the second half of the samples being obtained (15, 20, 25, 30, 40, 50, 60 min) at one time.

Bradford Analysis

250 μL samples from each timepoint were combined with 200 mL of 1:4 (Bradford:Buffer) Bradford reagent and 550 mL of additional Tris/CaCl2 buffer. UV-Vis spectra were obtained from 400-800 nm.

Bradford analysis new method.png

Proteinase K 1 uM New protocl.png