Smaller volumes of your sample will increase the resolution of your protein separation by size exclusion chromatography. Your sample should be highly concentrated. But be careful because at high concentrations your proteins might precipitate or viscosity effects can interfere with the separation. The sample volume can be expressed as a percentage of the total column volume. A typical recommendation for the sample volume is between 0.5 and 4 % of the column volume. In some cases the sample volume can be higher if the components that should be purified are well separated. For example, the sample volume for desalting columns is much higher than for classical SEC columns. The separation and the sample volume depend on the column and the sample composition and an optimal sample volume needs to be determined experimentally. If you have no experience with your sample and the column, we recommend starting with a sample volume of 1 % of the total column volume.