Objectives
- Prepare AuNP-protein solutions for biomineralization
- Analyze AuNP-protein solutions prepared on 10/30/2013 using AA spectroscopy.
AuNP solution preparation
- The general protocol for the preparation of AuNPs described on 08/28/2013 was followed.
- 20 AuNP solutions were prepared with BSA and Invertase proteins.
Table 1. Gold Standard Solution preparation
Preparation of Gold Stock Solution
|
''''
|
Molecular Weight (g/mol) |
393.83
|
mass Au(g) |
0.0354
|
volume stock(L) |
0.025
|
Concentration (M) |
0.00359546
|
Table 2. Preparation of BSA stock solution
Preparation of BSA stock solution
|
'
|
Molecular weight (g/mol) |
66000
|
mass (g) |
0.0432
|
volume stock (L) |
0.03
|
Concentration (M) |
2.18182E-05
|
Table 3. Preparation of Invertase Stock solution
Preparation Invertase Stock
|
'
|
Molecular weight (g/mol) |
135000
|
mass invertase (g) |
0.1015
|
volume stock (L) |
0.025
|
concentration (M) |
3.00741E-05
|
Table 4. Preparation of BSA-AuNP solutions
Ratio
|
Amount of gold added(mL)
|
concentration BSA (M)
|
volume BSA (mL)
|
volume Water added (mL)
|
105 |
0.347660664 |
2.38095E-06 |
0.545634921 |
4.106704416
|
110 |
0.347660664 |
2.27273E-06 |
0.520833333 |
4.131506003
|
115 |
0.347660664 |
2.17391E-06 |
0.498188406 |
4.15415093
|
120 |
0.347660664 |
2.08333E-06 |
0.477430556 |
4.174908781
|
125 |
0.347660664 |
0.000002 |
0.458333333 |
4.194006003
|
130 |
0.347660664 |
1.92308E-06 |
0.440705128 |
4.211634208
|
135 |
0.347660664 |
1.85185E-06 |
0.424382716 |
4.22795662
|
140 |
0.347660664 |
1.78571E-06 |
0.40922619 |
4.243113146
|
145 |
0.347660664 |
1.72414E-06 |
0.395114943 |
4.257224394
|
150 |
0.347660664 |
1.66667E-06 |
0.381944444 |
4.270394892
|
Table 5. Preparation of Invertase-AuNP solutions
Ratio
|
Amount gold added (mL)
|
Concentration Invertase (M)
|
Volume Invertase (mL)
|
Volume water added (mL)
|
30 |
0.347660664 |
8.33333E-06 |
1.38546798 |
3.266871356
|
50 |
0.347660664 |
0.000005 |
0.831280788 |
3.821058548
|
70 |
0.347660664 |
3.57143E-06 |
0.593771992 |
4.058567345
|
90 |
0.347660664 |
2.77778E-06 |
0.46182266 |
4.190516676
|
110 |
0.347660664 |
2.27273E-06 |
0.377854904 |
4.274484432
|
130 |
0.347660664 |
1.92308E-06 |
0.31972338 |
4.332615956
|
150 |
0.347660664 |
1.66667E-06 |
0.277093596 |
4.37524574
|
170 |
0.347660664 |
1.47059E-06 |
0.244494349 |
4.407844987
|
190 |
0.347660664 |
1.31579E-06 |
0.218758102 |
4.433581234
|
210 |
0.347660664 |
1.19048E-06 |
0.197923997 |
4.454415339
|
Notes
- All prepared protein-AuNP were colorless. However, the invertase-AuNP solutions prepared had a slightly cloudy appearance.
AA spectroscopy
General Protocol
- Transfer 3mL of sample from test tubes into a 10mL falcon tube.
- Centrifuge 3mL protein-AuNP solutions for 30 minutes at 4000rpm. Be sure that the centrifuge is balanced.
- Prepare 5 standard Au solutions (10μg/mL, 20μg/mL, 30μg/mL, 40μg/mL, 50μg/mL) following protocol described on 09/10/2013.
- Run samples on AA.
Table 6. Standard Au Solution preparation from 1000μg/mL AA Au gold standard solution
Concentration(μg/mL)
|
Volume of stock Au solution(mL)
|
Volume Deionized water added(mL)
|
10 |
0.1 |
9.9
|
20 |
0.2 |
9.8
|
30 |
0.3 |
9.7
|
40 |
0.4 |
9.6
|
50 |
0.5 |
9.5
|
Figure 1. Calibration curve of gold standard solutions
Figure 2. Atomic Absorbance Spectra of Myoglobin:AuNP solutions
Figure 3. Atomic Absorbance Spectra of Lysozyem:AuNP solutions
- The only solution that was colored was the 30 ratio. All other solutions appeared colorless.
Figure 4. Absorbance vs. Concentration plot of Myoglobin:AuNP solutions
Figure 5. Absorbance vs. Concentration plot of AuNP-Lysozyme Solutions
|