ABSTRACT
The belated acceptance of Micro-liquid chromatography (LC) contrasts sharply with the rapid proliferation of conventional high-performance liquid chromatography (HPLC) techniques in protein chemistry. Micro-LC techniques improve the sensitivity of peptide mapping by as much as twenty-fold, allowing multiple analytical runs to be made with smaller amounts of material. Two-dimensional HPLC purifications are thus becoming common where a crude protein extract or complex protein digest is fractionated on an ion-exchange column, followed by further purification of each fraction by injection and gradient elution on a reversed-phase column. The introduction of wide-pore supports, and the discovery of trifluoroacetic acid as a mobile phase modifier improved the efficiency and recovery of proteins to an extent that revolutionized protein purification technology. The most striking advantage of a Micro-LC is the higher sensitivity which enables purification of samples containing less than 10 nanogram of material.
