User:Michael F. Nagle/Notebook/Chem 571/2012/11/07: Difference between revisions
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==Objective== | |||
*Determine concentration of ADA prepared [[User:Michael_F._Nagle/Notebook/Chem_571/2012/10/23|Oct. 23]] via Bradford Assay | |||
==Bradford Assay== | |||
*Solutions were prepared with 715μL 1x Bio-Rad Bradford reagant and the following concentrations of BSA: | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''volume BSA (uL)''' | |||
| align="center" style="background:#f0f0f0;"|'''volume H2O (uL''' | |||
| align="center" style="background:#f0f0f0;"|'''[BSA] (mg/mL)''' | |||
| align="center" style="background:#f0f0f0;"|'''[BSA] (M)''' | |||
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| 0||30||0||0.00E+00 | |||
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| 5||25||0.24||3.61E-06 | |||
|- | |||
| 10||20||0.15||2.26E-06 | |||
|- | |||
| 15||15||0.13||2.01E-06 | |||
|- | |||
| 20||10||0.15||2.26E-06 | |||
|- | |||
| 25||5||0.24||3.61E-06 | |||
|- | |||
| 30||0||1||1.51E-05 | |||
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<Br> | |||
*A UV/Vis calibration curve was obtained by analyzing the samples and plotting BSA concentration against absorbance at 595nm. | |||
*The [[User:Michael_F._Nagle/Notebook/Chem_571/2012/11/06|ADA purified yesterday]], labelled Frac 9, was analyzed along with Fracs 2 and 8 purified [[User:Michael_F._Nagle/Notebook/Chem_571/2012/09/26|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]] | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Fraction #''' | |||
| align="center" style="background:#f0f0f0;"|'''Absorbance''' | |||
| align="center" style="background:#f0f0f0;"|'''[ADA] [mg/mL]''' | |||
| align="center" style="background:#f0f0f0;"|'''molar absorptivity [mL/mg*cm-1]''' | |||
| align="center" style="background:#f0f0f0;"|'''Molarity (µM)''' | |||
|- | |||
| 2||0.93||0.0177||52.4||42.0 | |||
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| 8||0.757||0.0115||65.9||54.4 | |||
|- | |||
| 9||0.819||0.0137||59.7||65.0 | |||
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<br>Frac 2 was found to be 42µM, while 8 was 54.4µM and 9 was 65µM. | |||
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Objective
Bradford Assay
Data
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