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(AuNP solution preparation)
(Objectives)
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#Analyze AuNP-protein solutions prepared on [[User:Moira M. Esson/Notebook/CHEM-571/2013/10/30|10/30/2013]] using AA spectroscopy.
 
#Analyze AuNP-protein solutions prepared on [[User:Moira M. Esson/Notebook/CHEM-571/2013/10/30|10/30/2013]] using AA spectroscopy.
 
<br>
 
<br>
 +
 
==AuNP solution preparation==
 
==AuNP solution preparation==
 
*The general protocol for the preparation of AuNPs described on [[User:Moira M. Esson/Notebook/CHEM-571/2013/08/28|08/28/2013]] was followed.
 
*The general protocol for the preparation of AuNPs described on [[User:Moira M. Esson/Notebook/CHEM-571/2013/08/28|08/28/2013]] was followed.
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<br>
 
<br>
 
{| {{table}}
 
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Molecular weight Au (g/mol)'''
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| align="center" style="background:#f0f0f0;"|''' Preparation of Gold Stock Solution'''
| align="center" style="background:#f0f0f0;"|'''393.83'''
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| align="center" style="background:#f0f0f0;"|'''''''''
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|-
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|Molecular Weight (g/mol)||393.83
 
|-
 
|-
 
| mass Au(g)||0.0354
 
| mass Au(g)||0.0354
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<br>
 
<br>
 
{| {{table}}
 
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Molecular weight (g/mol)'''
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| align="center" style="background:#f0f0f0;"|'''Preparation of BSA stock solution'''
| align="center" style="background:#f0f0f0;"|'''66000'''
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| align="center" style="background:#f0f0f0;"|''''''
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|-
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|Molecular weight (g/mol)||66000
 
|-
 
|-
 
| mass (g)||0.0432
 
| mass (g)||0.0432
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|}
 
|}
 
<br>
 
<br>
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==Notes==
 
==Notes==
 
*All prepared protein-AuNP were colorless. However, the invertase-AuNP solutions prepared had a slightly cloudy appearance.
 
*All prepared protein-AuNP were colorless. However, the invertase-AuNP solutions prepared had a slightly cloudy appearance.
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==AA spectroscopy==
 
==AA spectroscopy==
 
<br>
 
<br>
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'''General Protocol'''
 
#Transfer 3mL of sample from test tubes into a 10mL falcon tube.
 
#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.
 
#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 [[User:Moira M. Esson/Notebook/CHEM-571/2013/09/10|09/10/2013]].
 
#Prepare 5 standard Au solutions (10μg/mL, 20μg/mL, 30μg/mL, 40μg/mL, 50μg/mL) following protocol described on [[User:Moira M. Esson/Notebook/CHEM-571/2013/09/10|09/10/2013]].
 
#Run samples on AA.
 
#Run samples on AA.
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<br>
 +
Table 6. Standard Au Solution preparation from 1000μg/mL AA Au gold standard solution
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<br>
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{| {{table}}
 +
| align="center" style="background:#f0f0f0;"|'''Concentration(μg/mL)'''
 +
| align="center" style="background:#f0f0f0;"|'''Volume of stock Au solution(mL)'''
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| align="center" style="background:#f0f0f0;"|'''Volume Deionized water added(mL)'''
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|-
 +
| 10||0.1||9.9
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|-
 +
| 20||0.2||9.8
 +
|-
 +
| 30||0.3||9.7
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|-
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| 40||0.4||9.6
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|-
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| 50||0.5||9.5
 +
|}
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<br>
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Figure 1. Calibration curve of gold standard solutions
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<br>
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[[Image:Calibration Curve of Gold Standards zem11192013 .png]]
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<br>
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Figure 2. Atomic Absorbance Spectra of Myoglobin:AuNP solutions
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<br>
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[[Image:Atomic Absorbance Spectra of Myoglobin AuNP solutions zem11192013.png]]
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<br>
 +
Figure 3. Atomic Absorbance Spectra of Lysozyem:AuNP solutions
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<br>
 +
[[Image:Atomic Absorbance Spectra of Lysozyme AuNP solutions zem 11192013 .png]]
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<br>
 +
*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
 +
<br>
 +
[[Image:Absorbance vs. Concentration plot of Myoglobin-AuNP solutions zem11192013.png]]
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<br>
 +
Figure 5. Absorbance vs. Concentration plot of AuNP-Lysozyme Solutions
 +
<br>
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[[Image:Absorbance vs. Concentration plot of Lysozyme-AuNP solutions zem11192013.png]]
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 +
  
  

Revision as of 11:06, 10 December 2013

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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)
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

  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)
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
Calibration Curve of Gold Standards zem11192013 .png
Figure 2. Atomic Absorbance Spectra of Myoglobin:AuNP solutions
Atomic Absorbance Spectra of Myoglobin AuNP solutions zem11192013.png
Figure 3. Atomic Absorbance Spectra of Lysozyem:AuNP solutions
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
Absorbance vs. Concentration plot of Myoglobin-AuNP solutions zem11192013.png
Figure 5. Absorbance vs. Concentration plot of AuNP-Lysozyme Solutions
Absorbance vs. Concentration plot of Lysozyme-AuNP solutions zem11192013.png