Separation of intact proteins by using polyhedral oligomeric silsesquioxane based hybrid monolithic capillary columns.

High-efficient separation of intact proteins is still a huge challenge in proteome analysis of complex biological samples by using capillary columns. In this study, four POSS-based hybrid monolithic capillary columns were prepared and applied in nano-flow liquid chromatography (Nano-LC) separation of intact proteins. It was observed that the POSS-based hybrid monolithic columns exhibit high permeability, good LC separation reproducibility and column efficiency for intact protein separation. The effects of different LC separation conditions such as flow rate, gradient steepness, column length and mobile phase additives on the LC separation efficiency of the POSS-based hybrid monolithic column were systematically examined. Finally, fast LC separation of 7 proteins mixture was realized in 2.5 min by using the optimized conditions on the 100 μm i.d. POSS-based hybrid monolithic capillary column.