Achievable separation performance and analysis time in current liquid chromatographic practice for monoclonal...

The separation performance of a chromatographic system is often described in terms of column efficiency and peak capacity. Thanks to the new developments in column technology over the past few years, the achievable peak capacity drastically improved and the analysis time can be significantly shortened. Indeed, highly efficient wide-pore reversed-phase (RPLC) materials packed with small fully porous and superficially porous particles can be successfully used for the analytical characterization of therapeutic proteins. For non denaturating chromatographic approaches, such as ion exchange (IEX) and size-exclusion chromatography (SEC), non-porous ion-exchanger as well as sub -3μm size exclusion supports are commercially available and open new avenues in protein separations. In this study, the current possibilities offered by chromatography for the characterization of monoclonal antibody (mAb) are discussed. For this purpose, recently published data have been reviewed and calculations were performed to compare the maximum achievable peak capacity and related analysis times using typical samples under RPLC, IEX and SEC conditions. Carefully chosen realistic column pressure, mobile phase temperature, flow rate and column dimensions were considered for the case studies discussed through the paper.