User:Michael F. Nagle/Notebook/Chem 571/2012/11/07

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Objective

  • Determine concentration of ADA prepared Oct. 23 via Bradford Assay

Bradford Assay

  • Solutions were prepared with 715μL 1x Bio-Rad Bradford reagant and the following concentrations of BSA:
volume BSA (uL) volume H2O (uL [BSA] (mg/mL) [BSA] (M)
03000.00E+00
5250.243.61E-06
10200.152.26E-06
15150.132.01E-06
20100.152.26E-06
2550.243.61E-06
30011.51E-05


  • A UV/Vis calibration curve was obtained by analyzing the samples and plotting BSA concentration against absorbance at 595nm.
  • The ADA purified yesterday, labelled Frac 9, was analyzed along with Fracs 2 and 8 purified 9/26 via UV/Vis and it's peak was compared to the linear regression of BSA's in order to determine concentration.

Data

Image:Bradforassay.png

Fraction # Absorbance [ADA] [mg/mL] molar absorptivity [mL/mg*cm-1] Molarity (µM)
20.930.017752.442.0
80.7570.011565.954.4
90.8190.013759.765.0


Frac 2 was found to be 42µM, while 8 was 54.4µM and 9 was 65µM.


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