Fluorescence
Purpose
Use fluorescence to examine the rate at which myoglobin unfolds at pH4
Procedure
- Sample was made Myoglobin: 0.25mM Au, 12.5uM Myoglobin, 1.0mM HCl 3mL total
- Au 4.57mM
- (0.25mM)(3.0mL)=(XmL)(4.57mM)
- XmL= .176mL= 176uL Au
- Myoglobin 160uM
- (12.5uM)(3.0mL)=(XmL)(160uM)
- X=0.234mL= 234uL Myoglobin
- HCl 1.0mM
- (3.0mL)(1.0mM)=(1.0mM)(XmL)
- XmL=3.0mL= 300uL HCl
- Water
- Volume of water= 3000uL- 176uL-234uL-300uL= 2290uL
- Absorbance measurements were taken and saved every 3 minutes for 3 hours
- The instrument was set at 80 degrees Celsius
- The scan range parameters were start at 310nm and ending at 540nm
- The emission and exit slit were set at 10nm
Solutions for UV-VIS preparation
Stock Concentrations:
- Au 4.57mM
- Myoglobin 160mM
- Fructose 40.9mM
0.25 mM Fructose
pH
|
μL of 1 mM HClstock
|
μL of 1 mM NaOHstock
|
μL of 1 M NaOHstock
|
μL of AuCl3stock
|
μL of Fructosestock
|
μL of Myoglobinstock
|
μL of Water
|
4 |
500 |
0 |
0 |
293 |
31 |
388 |
3788
|
5 |
50 |
0 |
0 |
293 |
31 |
388 |
4238
|
6 |
5 |
0 |
0 |
293 |
31 |
388 |
4283
|
7 |
0 |
0 |
0 |
293 |
31 |
388 |
4288
|
8 |
0 |
5 |
0 |
293 |
31 |
388 |
4283
|
9 |
0 |
50 |
0 |
293 |
31 |
388 |
4238
|
10 |
0 |
500 |
0 |
293 |
31 |
388 |
3788
|
11 |
0 |
0 |
5 |
293 |
31 |
388 |
4283
|
12 |
0 |
0 |
50 |
293 |
31 |
388 |
4238
|
0.75 mM Fructose
pH
|
μL of 1 mM HClstock
|
μL of 1 mM NaOHstock
|
μL of 1 M NaOHstock
|
μL of AuCl3stock
|
μL of Fructosestock
|
μL of Myoglobinstock
|
μL of Water
|
4 |
500 |
0 |
0 |
293 |
92 |
388 |
3727
|
5 |
50 |
0 |
0 |
293 |
92 |
388 |
4177
|
6 |
5 |
0 |
0 |
293 |
92 |
388 |
4222
|
7 |
0 |
0 |
0 |
293 |
92 |
388 |
4227
|
8 |
0 |
5 |
0 |
293 |
92 |
388 |
4222
|
9 |
0 |
50 |
0 |
293 |
92 |
388 |
4177
|
10 |
0 |
500 |
0 |
293 |
92 |
388 |
3727
|
11 |
0 |
0 |
5 |
293 |
92 |
388 |
4222
|
12 |
0 |
0 |
50 |
293 |
92 |
388 |
4177
|
1.25 mM Fructose
pH
|
μL of 1 mM HClstock
|
μL of 1 mM NaOHstock
|
μL of 1 M NaOHstock
|
μL of AuCl3stock
|
μL of Fructosestock
|
μL of Myoglobinstock
|
μL of Water
|
4 |
500 |
0 |
0 |
293 |
159 |
388 |
3666
|
5 |
50 |
0 |
0 |
293 |
159 |
388 |
4116
|
6 |
5 |
0 |
0 |
293 |
159 |
388 |
4161
|
7 |
0 |
0 |
0 |
293 |
159 |
388 |
4166
|
8 |
0 |
5 |
0 |
293 |
159 |
388 |
4161
|
9 |
0 |
50 |
0 |
293 |
159 |
388 |
4116
|
10 |
0 |
500 |
0 |
293 |
159 |
388 |
3666
|
11 |
0 |
0 |
5 |
293 |
159 |
388 |
4161
|
12 |
0 |
0 |
50 |
293 |
159 |
388 |
4116
|
Results
After the sample was run through fluorescence, no color change had occurred indicating there was a source of error in the data collected, possibly from there being no gold in the sample prepared. The graphs for the incorrect data were still made and are shown below. The correct sample was run on 09/29/2016 by Matt Hartings and the correct graphs were created.
Graphs For Incorrect Data
Integrated intensity for Myoglobin templated Gold Nanoparticles at pH4
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Maximum intensity for Myoglobin templated Gold Nanoparticles at pH4
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Maximum Wavelength for Myoglobin templated Gold Nanoparticles at pH4
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- The graphs for the correct data were created by Matt Hartings. Data was collected and interpreted on 09/28/2016
Notes
pH4 was already done on 08/31/2016 but the prepared solution had incorrect concentrations of myoglobin and gold NPs
|