User:Monika Gasiorek/Notebook/CHEM-571 2014F/2014/11/05

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November 5, 2014

Primary Experiment #5 Work-up

30:1 colloid solution vs KI using MWCO 3500

Well 1 Well 2 Well 3 Well 4 Well 5
30:1 colloid 30:1 colloid 30:1 colloid 30:1 colloid 30:1 colloid
2 mM KI 5 mM KI 10 mM KI 25 mM KI 50 mM KI

Necessary Analyses:

Bradford

  • 5 colloid samples, 30:1 colloid reference, and Bradford blank
  • 200 μL sample, 200 μL 1:4 Bradford, 600 μL 50 mM Tris/50 mM NaCl
    • Results will be posted

Fluorescence & UV-VIS not necessary per NOTE on 10/21

I- ISE

  • 10 samples
Concentration Colloid Solution Potential (eV) Soak Solution Potential (eV)
50 mM-243.0-245.2
25 mM-225.2-222.5
10 mM-202.8-204.9
5 mM-186.9-187.9
2 mM-163.0-165.1

Primary Experiment #6 Set-up

0.6 g/L Lysozyme vs CaCl2 using MWCO 3500

Well 1 Well 2 Well 3 Well 4 Well 5
0.6 g/L lysozyme 0.6 g/L lysozyme 0.6 g/L lysozyme 0.6 g/L lysozyme 0.6 g/L lysozome
45 mM CaCl2 35 mM CaCl2 25 mM CaCl2 15 mM CaCl2 5 mM CaCl2

NOTE: Due to the results of previous experiments, it has been observed that the lysozyme (and colloid) precipitate out of solution at concentrations between 25 and 50 mM. Additionally, Ca2+ ISE measurements were unstable at concentrations below 0.5 mM. The Ca2+ concentration range has been adjusted accordingly.

Enzyme Kinetics

  • Kinetics mode on UV-VIS
  • 10mL of 400 unit/mL of lysozyme was prepared
    • Cold buffer was used
  • The procedure in the protocol was followed (see link in yesterday's tasklist)
  • The protocol was repeated using 4 solutions between 200 & 400 units/mL
    • Preparation was done just before addition to keep solution cold

NOTE: We never actually got to do this because the UV was not accessible...


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