Size exclusion chromatography loading capacity

Loading capacities in size exclusion chromatography are very low because all separation occurs within the liquid volume of the column. The small diffusion coefficients of macromolecules also contribute to bandspreading when loads are increased. The mass loading capacities for ovalbumin (MW 45,000) on various sizes of columns can be seen in Table 10.5. The maximum volume that can be injected in size exclusion chromatography before bandspreading occurs is about 2% of the liquid column volume. The maximum injection volumes for columns of different dimensions can also be seen in Table 10.5. 

In molecular exclusion chromatography, resolution is directly related to the sample loading volume, since the higher the sample volume, the higher the volume in which the protein is eluted. Usually, sample volume should be 0.5-5% of the total bed volume. For volumes below 0.5% the sample becomes too diluted, while volumes above 5% work well only for molecules with large differences in size. For desalting, due to the pronounced difference in the molecular size of the components, sample volumes can reach up to 30% of the column capacity without a significant decrease in resolution. 

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