Agilent Bio SEC columns are used to separate proteins, oligonucleotides, and other complex biopolymers. This application note demonstrates the effect of pore size on separations of proteins and antibody using Agilent Bio SEC-3 HPLC columns.
Size exclusion chromatography (SEC) separates biopolymers based on molecular size. This document evaluates how media pore size influences resolution when analyzing proteins, oligonucleotides, and other complex biopolymers.
The note presents experimental data using Agilent Bio SEC-3 columns with 100A, 150A, and 300A pore sizes to separate a standard protein mix and mouse IgG. Results indicate that smaller pore sizes exclude larger molecules, while larger pore sizes provide better resolution for distinguishing between monomers, dimers, and aggregates.
The 100A column is only suitable for proteins with molecular weights less than 100,000.
The flow rate used for the experiment is 0.35 mL/min.
The 300A column is the suitable choice for analyzing monoclonal antibodies and their aggregates.