What is chromatofocusing?

1 Answer
May 17, 2017

Chromatofocusing is a protein separation technique that allows resolution of single proteins and other ampholytes from a complex mixture according to differences in their isoelectric points.


This technique was introduced by Stuyterman and his colleagues between 1977 and 1981 .Chromatofocusing utilises ion exchange resins and is typically performed on a fast protein liquid chromatography.

It uses an ion exhange column packing and internally generated pH gradient that travels through the column as a retained front. The interaction between the pH gradient and amphoteric substances such as proteins causes these substances to exit the chromatographic column at characteristic locations in the effluent as focussed bands.

This gradient elution chromatographic technique is a powerful purification tool with respect to proteins, as it can resolve very similar species barely differing by 0.02 pH limits, that may not separate well using traditional ion exchange strategies.