How to use HPLC for protein purity detection and content determination?
First, gel permeation chromatography
Molecular weight determination: Several proteins with known molecular weights were determined by HPLC, and a standard curve corresponding to the adjusted retention time was plotted. Then, the retention time of the unknown protein was measured under the same conditions, and the molecular weight of the unknown protein was compared with the standard curve. ;
Determination of content: Prepare a series of different concentrations of protein solution with protein standards, draw a standard curve of protein concentration and peak height/peak area, and then measure the peak height/peak area of the position sample under the same conditions to obtain an unknown sample. concentration;
Purity determination: The purity of the target protein can be obtained by dividing the peak area/peak height of the target protein by the sum of all peak areas/peak heights in the case where all components in the sample to be tested are peaked.