Team:UCLA/Notebook/Protein Cages/19 August 2015
Phillip's notes:
Introduction: BCA assay will be done on the final elution of the previous affinity purification. According to Kevin from the Yeate’s lab, a minimum of 2mg/mL of protein is needed for the DLS assay.
Procedures: The standard procedure was followed in the Thermo BCA kit in order to prepare the standard solutions. All standards were diluted in triton lysis buffer.
Elution 3 was used because it appeared to be the purest. Upon starting the assay, it was noted that some of the protein was aggregating and precipitating out of solution. The tube was spun down at max speed for 5 minutes, and the supernatant collected. The samples ran were the following:
Elution with aggregate, supernatant, 1:10 supernatant, and 1:100 supernatant.
A standard curve was constructed on excel. Only absorbance readings at about .1 to 1 were used. Only the original elution showed a measured protein concentration at about 620ug/mL.
Conclusion: It appears that most of the protein cage is aggregating out of solution. Perhaps this can be used in order to separate the protein cage in the other two elutions, and resuspend them in a higher volume.