User:Andrew W Long/Notebook/CHEM-671 Research/2015/09/29

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Bradford Assay with Proteinase-K & Fibers Over Time

Today a Bradford Assay was done on the products of a reaction between Proteinase-K and previously synthesized AuNP fibers. Using UV-Vis, samples were analyzed at 10min, 15min, 30min, 45min, 60min, 1.5hrs, and2hrs. An additional sample without AuNP fibers was analyzed as a control.


The concentration of a previously pepped and frozen stock sample (sample 2) of proteinase-K was calculated in the following manner: Failed to parse (MathML with SVG or PNG fallback (recommended for modern browsers and accessibility tools): Invalid response ("Math extension cannot connect to Restbase.") from server "https://api.formulasearchengine.com/v1/":): {\displaystyle \frac{0.00087gProteinaseK}{1}|\frac{1mol}{28,900g}=3.0104*10^-8mol} Failed to parse (MathML with SVG or PNG fallback (recommended for modern browsers and accessibility tools): Invalid response ("Math extension cannot connect to Restbase.") from server "https://api.formulasearchengine.com/v1/":): {\displaystyle \frac{3.0104*10^-8molProteinaseK}{0.001L}|\frac{L}{1000mL}=3.0104*10^-5M=30.1040 uM proteinase-K}


With the concentration of the proteinase-K solution, and with the knowledge that we need 1ml of a 1uM version of the solution, the needed volume of proteinase-K was calculated in the following manner: M1*V1=M2*V2

(30.1040uM)(V2)=(1uM)(1mL) ==> V1=0.0332mL = 33.2181 uL

(1ml - 33.2181 uL, so 966.8uL) of buffer was added to each of the 7 dried samples of protein fibers and vortexed. 0.0332uL of the proteinase-K solution was then added to each of the protein solutions, were allowed to incubate in a warm water bath for their respective times, and then were examined in the UV-VIS (i.e. after 10min, the 10min sample was examined in the UV-Vis). A graph displaying the results of the assay is shown below:

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The procedure can be found here in Dr. Harting's Notebook.