GE AKTA avant User Manual page 224

5 Operation
5.8 Fractionation
5.8.3 Fractionation overview
Peak broadening
The width of peaks at the fraction collector is influenced by the properties of the column
and the dimensions of tubing connecting the components and the components them-
selves. Initial sample volume affects the peak width in gel filtration (GF) chromatography.
A sample zone is broadened during passage through a GF column so that the sample
is diluted and the resolution decreases with increasing sample volume. Sample volume
does not however affect the resolution in adsorption chromatography techniques such
as affinity chromatography (AC), ion exchange chromatography (IEX), and hydrophobic
interaction chromatography (HIC). The effect of peak broadening in the system from
sample injection to peak detection (including dilution on the column) is apparent in the
chromatogram from the UV monitor, but broadening from the UV monitor to fraction
collection is not visible in the chromatogram. With small analytical columns the peak
broadening between the UV monitor and the fraction collector will be obvious.
224 ÄKTA avant User Manual 29035184 AE