User:Andrew W Long/Notebook/CHEM-671 Research/2015/10/27: Difference between revisions
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==Bradford Assay, 10nM== | ==Bradford Assay, 10nM== | ||
The instructions for the day can be found [[User:Matt_Hartings/Notebook/AU_Biomaterials_Design_Lab/2015/10/06|here]] | |||
[[Image: | Previously synthesized fiber samples were prepared in the following way. First, 7 fiber samples were centrifuged at 300rpm for 10min, and the supernatant was removed. Stock of proteinase-K was suspended with 1mL of Tris buffer (50mM) with CaCl (10mM) at pH 8. The concentration of proteinase-K of the resulting solution was then determined to be 41868 μM. This was calculated using (0.00121g)(1mol/28900g)(1/0.001L). This solution was diluted in the following way to get a concentration of 0.238843 uL. (41868 nM)*(V1) = (10nM)*(1mL). The amount of buffer required to make the solution 1mL was determined to be 976.1uL to make 1mL of sample. Next, the 7 samples were incubated in a shaking hot water bath, as well as the 7 blanks. Once each sample had incubated for it's allotted time, it was pulled out of the bath, and centrifuged at 300rpm for 1min. 750mL was placed in a plastic cuvette, with 600mL dilute Bradford stock solution, and 1650uL buffer. The time periods were 10min, 15min, 30min, 45min, 1hr, 1.5hr, and 2hr. The samples were scanned from 400-800nm using the UV-Vis. The results from the experiment are shown below: | ||
[[Image:Bradford_Assay_of_10nM_AuNP_Fibers_with_Lysozyme_Digestion%2C_Wavelength_vs._Absorbance_at_Various_Times_.png|600px]] | |||
[[Image:AMS_Bradford_Assay_of_10nM_AuNP_Fibers_with_Lysozyme_Digestion%2C_Absorbance_at_600nm_vs._Time.png|600px]] | |||
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Bradford Assay, 10nMThe instructions for the day can be found here Previously synthesized fiber samples were prepared in the following way. First, 7 fiber samples were centrifuged at 300rpm for 10min, and the supernatant was removed. Stock of proteinase-K was suspended with 1mL of Tris buffer (50mM) with CaCl (10mM) at pH 8. The concentration of proteinase-K of the resulting solution was then determined to be 41868 μM. This was calculated using (0.00121g)(1mol/28900g)(1/0.001L). This solution was diluted in the following way to get a concentration of 0.238843 uL. (41868 nM)*(V1) = (10nM)*(1mL). The amount of buffer required to make the solution 1mL was determined to be 976.1uL to make 1mL of sample. Next, the 7 samples were incubated in a shaking hot water bath, as well as the 7 blanks. Once each sample had incubated for it's allotted time, it was pulled out of the bath, and centrifuged at 300rpm for 1min. 750mL was placed in a plastic cuvette, with 600mL dilute Bradford stock solution, and 1650uL buffer. The time periods were 10min, 15min, 30min, 45min, 1hr, 1.5hr, and 2hr. The samples were scanned from 400-800nm using the UV-Vis. The results from the experiment are shown below:
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