User:Monika Gasiorek/Notebook/CHEM-571 2014F/2014/09/24

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September 24, 2014

Bradford Analysis of Dialysis

A Bradford analysis was done on the dialysis samples prepared and dialyzed last week.

  • 600 µL of each soak solution were combined with 200 µL of Bradford reagent and 200 µL of a 50 mM Tris/NaCl buffer
    • The soak solutions included:
                              50 mM NaCl stock solution with 3500 g/mol MWCO tubing
                              50 mM Glycine stock solution with 3500 g/mol MWCO tubing
                              Pure water with 3500 g/mol MWCO tubing 
                              50 mM Glycine stock solution with 25k g/mol MWCO tubing
                              50 mM Glycine stock solution with 3500 g/mol MWCO tubing
  • 100 µL of the contents of each dialysis tubing was combined with 200 µL of Bradford reagent and 700 µL of Tris/NaCl buffer
    • The respective dialysis bag contents were analyzed, all of which initially included 4 mL of 1g/L lysozyme stock solution, except for the very last system, in which 4.0 mL of 36.5 [Au]:[Lys] colloid solution was used in the place of lysozyme against the 50 mM glycine soak with 3500 tubing

NOTE: As mentioned previously, the contents of the tube in the fourth system (50 mM glycine solution with 25K g/mol MWCO tubing) were lost during transfer; thus, only four tubing samples are included in the results.

Solution Conc of Lysozyme (µg/mL) Abs
3500 NaCl Soak-3.3446601940.028
3500 H2O Soak-3.9757281550.015
3500 Gly Soak2.5291262140.149
25k Gly Soak-0.092233010.095
3500 Gly Colloid1.7038834950.132
3500 NaCl/Lys Bag17.917475730.466
3500 H2O/ Lys Bag18.548543690.479
3500 Gly/Lys Bag20.053398060.51
3500 Gly/Coll Bag2.820388350.155

NOTE: Negative concentrations of protein in the soak solutions may be a result of protein precipitation.

CaCl2 Conductivity

A hard water probe sensitive only to specific polyvalent cations such as Ca2+ was used to measure the conductivity in mV of 5 solutions of CaCl2 of various concentrations.

  • 0.11098 g of CaCl2 was dissolved in 20 mL of water, making a 50 mM CaCl2 solution. 2 mL of this solution was then diluted with 18 mL of water to make the next serial dilution. This step was repeated 3 more times.

Concentration Conductivity (mV)
5 µM (-4.1)
50 µM 7.3
0.5 mM 36.1
5 mM 63.6
50 mM 87.8

NOTE: The results were graphed with conductivity as a function of the log of CaCl2 concentration.


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