User:Moira M. Esson/Notebook/CHEM-571/2013/11/19

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Objectives

  1. Prepare AuNP-protein solutions for biomineralization
  2. 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)
1050.3476606642.38095E-060.5456349214.106704416
1100.3476606642.27273E-060.5208333334.131506003
1150.3476606642.17391E-060.4981884064.15415093
1200.3476606642.08333E-060.4774305564.174908781
1250.3476606640.0000020.4583333334.194006003
1300.3476606641.92308E-060.4407051284.211634208
1350.3476606641.85185E-060.4243827164.22795662
1400.3476606641.78571E-060.409226194.243113146
1450.3476606641.72414E-060.3951149434.257224394
1500.3476606641.66667E-060.3819444444.270394892


Table 5. Preparation of Invertase-AuNP solutions

Ratio Amount gold added (mL) Concentration Invertase (M) Volume Invertase (mL) Volume water added (mL)
300.3476606648.33333E-061.385467983.266871356
500.3476606640.0000050.8312807883.821058548
700.3476606643.57143E-060.5937719924.058567345
900.3476606642.77778E-060.461822664.190516676
1100.3476606642.27273E-060.3778549044.274484432
1300.3476606641.92308E-060.319723384.332615956
1500.3476606641.66667E-060.2770935964.37524574
1700.3476606641.47059E-060.2444943494.407844987
1900.3476606641.31579E-060.2187581024.433581234
2100.3476606641.19048E-060.1979239974.454415339


Notes

  • All prepared protein-AuNP were colorless. However, the invertase-AuNP solutions prepared had a slightly cloudy appearance.

AA spectroscopy


General Protocol

  1. Transfer 3mL of sample from test tubes into a 10mL falcon tube.
  2. Centrifuge 3mL protein-AuNP solutions for 30 minutes at 4000rpm. Be sure that the centrifuge is balanced.
  3. Prepare 5 standard Au solutions (10μg/mL, 20μg/mL, 30μg/mL, 40μg/mL, 50μg/mL) following protocol described on 09/10/2013.
  4. 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)
100.19.9
200.29.8
300.39.7
400.49.6
500.59.5


Figure 1. Calibration curve of gold standard solutions
Image:Calibration Curve of Gold Standards zem11192013 .png
Figure 2. Atomic Absorbance Spectra of Myoglobin:AuNP solutions
Image:Atomic Absorbance Spectra of Myoglobin AuNP solutions zem11192013.png
Figure 3. Atomic Absorbance Spectra of Lysozyem:AuNP solutions
Image:Atomic Absorbance Spectra of Lysozyme AuNP solutions zem 11192013 .png

  • 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
Image:Absorbance vs. Concentration plot of Myoglobin-AuNP solutions zem11192013.png
Figure 5. Absorbance vs. Concentration plot of AuNP-Lysozyme Solutions
Image:Absorbance vs. Concentration plot of Lysozyme-AuNP solutions zem11192013.png




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