2-DIMENSIONAL GEL-ELECTROPHORESIS LIQUID-CHROMATOGRAPHY FOR THE MICROPREPARATIVE ISOLATION OF PROTEINS

Although comprehensive, column-based two-dimensional separation techni ques offer enormous resolving power for a complex mixture, they often lack the ability to isolate the separated species for further analysis (e.g., proteins for Edman sequencing). This paper describes a micropr eparative two-dimensional separation system for the isolation of prote ins from complex mixtures, such as cell lysates. The system is compose d of commercially available equipment: continuous-elution tube gel ele ctrophoresis as the first dimension followed by gradient elution, reve rsed-phase perfusion liquid chromatography as the second dimension wit h a two-loop sampling valve as the interface between dimensions. The t wo-dimensional electrophoresis/liquid chromatography system (2D-EP/LC) shows high resolution of proteins since each dimension has orthogonal separation mechanisms (electrophoresis, size/charge; LC, hydrophobici ty). Identification of proteins for further analysis is accomplished b y superimposing a grid on the computer-generated 3D image.