Separation of proteins by microcolumn liquid chromatography based on the reversed-phase and size-exclusion principles

Slurry-packed fused-silica microcolumns of 250 micron I.D., are characterized for use in high-performance liquid chromatographic studies of proteins. The present work utilizes the reversed-phase and size-exclusion chromatographic modes for the separation of standard protein mixtures. A 5-micron, 300-A octyl material is utilized for the reversed-phase studies, and the size-exclusion studies are accomplished with 5-micron diol material of 60-, 100- and 300-A pore sizes. Column efficiency and packing reproducibility, as well as sample capacity in a micropreparative mode, are discussed. In addition, the inherent mass sensitivity of a microcolumn liquid chromatography system as applied to protein detection is demonstrated.