ABSTRACT
Liquid chromatography has been a very useful tool to biochemists for several decades. In the past few years, there have been an increasing number of papers showing examples of very fast separations of proteins by reversed-phase chromatography (RPC) and ion-exchange chromatography (IEC) using both nonporous and macroporous supports. This chapter describes a new tool, the Ultrafast Microprotein Analyzer (UMA), which allows rapid gradient separations by both RPC and IEC. The UMA has been specifically designed to allow rapid re-equilibration by minimizing volumes and optimizing flow dynamics, and uses a variety of new fast flow high-performance liquid chromatography packings in a 50 × 1 mm internal diameters column at 5 to 10 times the normal flow velocities for these microbore columns with standard packing materials. This allows highly efficient, reproducible separations with a total analysis time of under 5 min. The chapter also demonstrates the application of the UMA to ultrafast analysis of recombinant proteins by RPC.
