Size exclusion chromatography (SEC) is a noninteractive technique that separates solutes according to their molecular size in non-denaturing conditions. It is often used as the first step in isolation of a protein from a crude sample. We will discuss the theory behind SEC, overview of mAbs, and characterization requirements; with a focus specifically on the effect of solute size and molecular weight, general rules for using SEC, column selection choices, important mobile phase considerations, and column care. We will explore method development pointers in the troubleshooting section.