User:Puja Mody/Notebook/Chem 571: Gold Nanoparticles/2012/11/07

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(Procedure)
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* The absorbance values at 595 nm of each solution were graphed versus BSA concentration in mg/mL to produce a calibration curve.
* The absorbance values at 595 nm of each solution were graphed versus BSA concentration in mg/mL to produce a calibration curve.
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* Using linear regression, the equation of the line was obtained.
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==DATA==
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[[Image:Bradford_assay.png]]
[[Image:Bradford_assay.png]]
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* Fractions of ADA, made on [[User:Melissa_Novy/Notebook/CHEM-571/2012/10/03|2012/10/03]] and [[User:Melissa_Novy/Notebook/CHEM-571/2012/11/06|2012/11/06]], were obtained.
 
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* 7.5 μL of each fraction was added to 7.5 μL of autoclaved water and 715 μL of 1X Bradford reagent.
 
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** Fraction #2, obtained in November, was diluted by half, using the same procedure as listed above for the standard solutions, such that its absorbance was below 1.
 
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{| {{table}}
 
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| align="center" style="background:#f0f0f0;"|'''Sample Name'''
 
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| align="center" style="background:#f0f0f0;"|'''Volume of ADA [mL]'''
 
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| align="center" style="background:#f0f0f0;"|'''Absorbance []'''
 
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| align="center" style="background:#f0f0f0;"|'''Conc. ADA [mg/mL]'''
 
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| align="center" style="background:#f0f0f0;"|'''Mass of ADA [mg]'''
 
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| align="center" style="background:#f0f0f0;"|'''Moles of ADA [mol]'''
 
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| align="center" style="background:#f0f0f0;"|'''Molar Absorptivity of ADA [mL/mg cm]'''
 
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| align="center" style="background:#f0f0f0;"|'''Molarity [M]'''
 
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* When these samples were originally run, they produced absorbances over 1 which resulted in us needing to dilute the solution further. As a result, all solutions were diluted by a half to 715μL of original solution with an additional 715μL of water.
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* Linear regression was used to determine the equation of the line
 +
 
 +
* When these samples were originally run, they produced absorbances over 1 which resulted in us needing to dilute the solution further. As a result, all solutions were diluted by a half to 715μL of original solution with an additional 715μL of water.
==DATA==
==DATA==

Revision as of 16:41, 11 November 2012

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Bradford Assay

Goals

Procedure

  • A stock solution of 1mg/mL BSA was prepared by adding 10mg of BSA into a 10mL volumetric flask and then filling it with autoclaved water. The actual concentration of the BSA stock was 1.02mg/mL as the actual weighed out BSA added was 10.2mg BSA.
  • In order to determine the concentration of the ADA collected in the fracs, standard solutions had to be made and run through the UV-Vis first. Seven standard solutions were prepared with 1X Bradford Reagent, varying amounts of BSA stock, and autoclaved water. These solutions were analyzed in the UV-Vis from 500-800 nm.
Volume 1X Bradford Reagent Added [mL] Volume BSA Stock Added [mL] Volume Water Added [mL] Conc. BSA [mg/mL]
1.400.030
1.40.0050.0250.003566434
1.40.010.020.007132867
1.40.0150.0150.010699301
1.40.020.010.014265734
1.40.0250.0050.017832168
1.40.0300.021398601


  • The absorbance values at 595 nm of each solution were graphed versus BSA concentration in mg/mL to produce a calibration curve.

DATA

Image:Bradford_assay.png

  • Linear regression was used to determine the equation of the line


  • When these samples were originally run, they produced absorbances over 1 which resulted in us needing to dilute the solution further. As a result, all solutions were diluted by a half to 715μL of original solution with an additional 715μL of water.

DATA


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