User:Paul Rothenberg/Notebook/CHEM 571 Fall 2014/2014/10/14

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SDS-PAGE Prep

SDS-PAGE was prepared according to the following:

  • 10 μL 0.6 g/L lysozyme with 10μL SDS-PAGE sample buffer in 1.5 mL centrifuge vial
  • 10 μL 0.12 g/L lysozyme with 10μL SDS-PAGE sample buffer in 1.5 mL centrifuge vial
  • 10 μL 30:1 Au/lysozyme colloid with 10μL SDS-PAGE sample buffer in 1.5 mL centrifuge vial
  • 10 μL 0.12 g/L unknown protein with 10 μL SDS-PAGE sample buffer in 1.5 mL centrifuge vial
  • Placed in heating block (set at 90 °C) for 5 minutes, and placed in the fridge overnight.


A pH 6.24 66 mM Potassium Phosphate buffer was prepared using monobasic and dibasic solids.


Dialysis Analysis

Bradford, UV-VIS, and Fluorescence analysis were run on the dialysis samples from the KI dialysis. 1 in 4 Bradford solution was prepared and the samples were prepped the same method as previously. A blank was also run. For UV-VIS and fluorescence, a 1 in 100 dilution of each was prepared and run. UV-VIS was run from 200-400 nm.


Bradford Results

Bradford Lysozyme KI 10 14 MGPR.png

Wavelength Abs at 577 nm Concentration (ug/mL)
Stock 0.121 1.169902913
2 mM 0.115 0.878640777
5 mK 0.124 1.315533981
10 mM 0.114 0.830097087
25 mM 0.111 0.684466019
50 mM 0.115 0.878640777

Fluorescence

Lysozyme KI 3500 Fluorescence Emission Chart.png

UV-VIS

Lysozyme KI 3500MW UV VIS 1in100 Chart.png