User:Monika Gasiorek/Notebook/CHEM-571 2014F/2014/09/16

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September 16, 2014

Stock Solution Preparation

A 500 mL stock solution of 50 mM NaCl was made.

  • 1.461 g of solid NaCl was dissolved in a 500 mL volumetric flask

NOTE: The purpose of this solution was to serve as a dialyzing agent

A 1 L stock buffer solution of 50 mM glycine with a pH of approx. 3.5 (pKa in glycine buffer range) was made.

  • 3.7576 g of glycine was dissolved in a 1 L volumetric flask with the addition of 4.00 mL of 1 M HCl

NOTE: The purpose of this solution was also to serve as a dialyzing agent. A buffer solution was necessary in order to change the ionic strength of the solution without altering its pH.

    • Pure water results in the precipitation of the AuNP colloid; an acidic buffer solution is conducive to maintaining the desired structure in a desired pH range while allowing for changes in ionic strength/conductivity and the movement of ions across the dialysis tubing

A 25 mL solution of 1 g/L lysozyme was made.

  • 0.025 g of lysozyme was dissolved in 25.0 mL of water

NOTE: The purpose of this solution was to serve as a baseline for the measurement of the equilibria/movement of free protein and/or ions across the dialysis tubing.

A 10.0 mL sample of approximately 30:1 (rather 36.5:1) gold:protein colloid solution was obtained from previous experiments.

  • The sample came from the addition of 1 mL of 70.96μm lysozyme and 1 mL of 2.53 mM gold, and 8 mL of water in day one of lab - not included in lab notebook.

Dialysis

5 dialysis experiments were set up:

I - 4.0 mL of the prepared lysozyme solution was dialyzed in about 3 in of 3500 g/mol MWCO tubing in a 150 mL beaker filled with approx. 120 mL of the 50 mM NaCl stock solution.

II - 4.0 mL of the prepared lysozyme solution was dialyzed in about 3 in of 3500 g/mol MWCO tubing in a 150 mL beaker filled with approx. 120 mL of the 50 mM glycine stock solution.

III - 4.0 mL of the prepared lysozyme solution was dialyzed in about 3 in of 3500 g/mol MWCO tubing in a 150 mL beaker filled with approx. 120 mL of pure water.

IV - 4.0 mL of the prepared lysozyme solution was dialyzed in about 3 in of 25k g/mol MWCO tubing in a 150 mL beaker filled with approx. 120 mL of the 50 mM glycine stock solution.

V - 4.0 mL of the 35.6:1 [Au]:[lys] solution wad dialyzed in about 3 in of 3500 g/mol MWCO tubing in a 150 mL beaker filled with approx. 120 mL of the 50 mM glycine stock solution.

NOTE: A sixth dialysis using 15k g/mol MWCO tubing was intended (4.0 mL of lysozyme solution against 50 mM glycine soln), but the specific tubing was not available.

Calibration Curves: pH and Conductivity

pH and conductivity measurements were obtained for the baseline, as well as 4 serial dilutions (2x, 5x, 10x, 20x) of the 50 mM NaCl solution.

Concentration (mM) pH Conductivity (μS/cm)
50 8.827 5.56 (mS/cm)
25 8.357 2924
10 8.199 1173
5 8.075 638
2.5 7.873 315

NOTE: To be continued...



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