Objective
- To prepare gold fibers that we'll will use for the rest of the semester.
- To prepare protease samples that we will use for the rest of the semester and record the mass of 15 eppendorf tubes.
- To compile the class's UV/Vis, florescence, and average eppendorf tube mass data.
- To run our Au fibers with trypsin on the Ocean Optics Spec.
Procedures
- Preparing gold fiber
- We followed the protocol from September 2nd except we used the stock concentration made by Dr. Hartings today, see description.
- Preparing protease samples
- We weighed and recorded the mass of an eppendorf tube.
- We tared the scale and added between 0.001 g to 0.002 g of protease to the tube.
- We labeled the tube with the date, molarity (if 1 mL of water is added), protease (Trypsin), and RAM.
- We repeated steps 1-3 for to create 15 protease samples and stored in our freezer box.
- Last we uploaded the weights of the 15 eppendorf tubes to dropbox, and averaged all the groups weights.
Ocean Optics
Instrument settings - total measurement for 2.5 hours and a reading was recorded ever 2 minutes.
Sample preparation
- We prepared a 1 mL sample of 52.36 µM of Trypsin and water solution. Then we calculated sample solution we needed to make a 1µM sample in 3 mL.
- We added 1 mL of gold fibers, 0.3 mL of buffer, and 0.0573 mL of trypsin after the second reading.
- We added 1.643 mL of water to reach 3 mL of sample in the cuvette.
Calculations for volume of trypsin:
C1·V1 = C2·V2,
where C1 = 52.36µM
C2 = 1µM
V2 = 3mL
V1 = 57.3µL
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
- HAuCl4·H2O
- 2.5 mM
- MW = 339.79 g/mol
- Volume = 50 mL
- 42.4 mg
- Lysozyme
- 20 μM
- MW = 14307 g/mol
- Volume = 100 mL
- 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
- HAuCl4·H2O
- 50.38 mg
- MW = 339.79 g/mol
- Volume = 50 mL
- 2.96 mM
- Lysozyme
- 28.69 mg
- MW = 14307 g/mol
- Volume = 100 mL
- 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 mL samples
- Volume of gold stock: 84.4 μL
- Volume of protein stock: 276 μL
- Volume of water: 640 μL
- 5 mL samples
- Volume of gold stock: 422 μL
- Volume of protein stock: 1,380 μL
- Volume of water: 3,198 μL
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
|
- Ocean Optics graphs are below
Results
- We did not create any gold nanoparticles or nanofibers.
- The average mass of an eppendorf tube is 1.025135426 and the standard deviation was 0.005373647.
- As a class the Lysozyme Molar Absorbtivity Coefficient: 33200 M-1 cm-1 and an R2 equal to 0.9846.
- INSERT OCEAN OPTICS RESULTS HERE
Matt HartingsYou need to discuss the results from #1, #2, and #3 from your objectives. What is the data from the tube measurement? What was the result of your measurements with the ocean optics? What were the results of your fiber synthesis?
|