User:M Jaffe/Notebook/Maxis Protease Lab 471/2015/09/15

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Objective

We made more fibers for analysis to use for the rest of the semester.

  1. We'll be using the protocols we used on September 2nd. The only difference is that we'll use the stock concentrations that Professor Hartings made today (see below).

Description

Target Concentrations

Based on the total reaction volume that the class will be using and the final concentrations that will be in each reaction volume, I have determined that the following HAuCl4·H2O and Lysozyme masses are appropriate for today's stock solutions.

Stock solutions

  1. HAuCl4·H2O
    1. 2.5 mM
    2. MW = 339.79 g/mol
    3. Volume = 50 mL
    4. 42.4 mg
  2. Lysozyme
    1. 20 μM
    2. MW = 14307 g/mol
    3. Volume = 100 mL
    4. 28.6 mg

Actual Concentrations

The actual masses are different from the target masses. The concentrations shown below reflect the actual concentrations made for the stock solutions. I used a 50 mL volumetric flask to make the gold stock solution and a 100 mL volumetric flask to make the protein solution. In each case I used HPLC-grade water.

Stock solutions

  1. HAuCl4·H2O
    1. 50.38 mg
    2. MW = 339.79 g/mol
    3. Volume = 50 mL
    4. 2.96 mM
  2. Lysozyme
    1. 28.69 mg
    2. MW = 14307 g/mol
    3. Volume = 100 mL
    4. 20.1 μM

Directions for making samples

Note: The final concentration of gold should be 0.25 mM and the final concentration of lysozyme should be 5.556 μM. This gives a Au:lysozyme ratio of 45, which should yield fibers.

  1. 1 mL samples
    1. Volume of gold stock: 84.4 μL
    2. Volume of protein stock: 276 μL
    3. Volume of water: 640 μL
  2. 5 mL samples
    1. Volume of gold stock: 422 μL
    2. Volume of protein stock: 1,380 μL
    3. Volume of water: 3,198 μL


  • Preparation of trypsin samples.
  1. Weigh 15 eppendorf tubes and record their mass.
  2. To each tube, add approximately 1mg of trypsin, weigh and record.
  3. Calculate the concentration in each tube if 1mL of liquid were added.

Calculations to find concentration:

    Concentration = [trypsin(g)/23'300g/mol]/0.001L


Matt Hartings What were the results of your syntheses? Where is the class UV-Vis analysis? What is the eppendorf mass data?

Data

The table below lists the protease samples we prepared and the mass of the corresponding eppendorf tube.

Mass of Eppy Tube (g) Mass of Protease (g) Concentraton (micromolar)
1.02691 0.00108 46.35
1.02622 0.00148 63.52
1.0224 0.00117 50.21
1.01944 0.00169 72.33
1.02922 0.00181 77.68
1.02625 0.00128 54.93
1.01909 0.00165 70.82
1.02036 0.00182 78.11
1.01922 0.00128 54.93
1.02024 0.00122 52.36
1.02097 0.00154 66.09
1.02422 0.00175 75.11
1.02669 0.00124 53.22
1.02936 0.00148 63.52
1.02923 0.00155 66.52

The table below lists the the class's eppendof tube masses (my groups data set are under 'RAM 9/15').

AMS 9/9 AMS 9/15 LMN 9/9 JBN 9/9 MB 9/9 RAM 9/15 AMS 9/16
1.02157 1.03107 1.03104 1.015 1.026 1.03005 1.03028
1.03162 1.03192 1.02859 1.01348 1.026 1.0255 1.02355
1.02798 1.02697 1.0199 1.0179 1.026 1.02676 1.02919
1.03275 1.01923 1.02753 1.02954 1.026 1.02869 1.02559
1.03057 1.02998 1.01646 1.0228 1.026 1.01656
1.01392 1.03059 1.03078 1.02048 1.026 1.01995
1.02938 1.02025 1.02419 1.01945 1.026 1.01707
1.02917 1.02932 1.01308 1.02723 1.026 1.02417
1.02993 1.03331 1.02896 1.02951 1.026 1.02518
1.02223 1.0287 1.0288 1.01517 1.026 1.01762
1.02055 1.02554 1.02602 1.02954 1.026 1.0349
1.03129 1.01372 1.02456 1.03211 1.026 1.03024
1.01359 1.01853 1.01719 1.0192 1.026 1.02016
1.02997 1.02051 1.02863 1.02781 1.026 1.0205
1.03117 1.03015 1.0285 1.026 1.02812 1.01972
Average Mass (g) 1.025135426 Standard Deviation 0.005373647

The table below is the raw data for the graph following it. The graph is a concentration vs absorbance of the class's compiled UV/Vis data at 280 nm.

Group Concentration A(280nm)
Group 1 2 0.052
5 0.16
8 0.239
10 0.292
15 0.444
Group 3 0.7875 0.026
2.625 0.091
5.25 0.186
7.5 0.252
15 0.502
Group 4 0.965 0.032
1.93 0.069
3.86 0.123
7.225 0.261
15.445 0.517
Group 5 1.56 0.052
3.13 0.108
6.25 0.221
12.5 0.441
15 0.54


571ClassesUVVISGRAPHMABC.png