ABSTRACT
The goal of this book has been to present the broad spectrum of separation methods for protein analysis by capillary electrophoresis and to provide some guidelines in their practical application. These methods include separations based on mass and charge, isoelectric point, molecular sieving, and affinity interactions. In fact, CE separation modes analogous to most existing electrophoretic and some chromatographic techniques are currently available. In many cases, resolving power exceeds that of conventional methods by virtue of the excellent heat dissipation of fused silica capillaries and the plug-flow characteristics of EOF. In spite of these advantages, CE is now only beginning to be established as a routine analytical tool in the protein chemistry lab. This can be attributed to limitations in reproducibility, sensitivity, and throughput. It seems appropriate at the conclusion of this work to address these issues with an eye to the future evolution of protein CEo
Reproducibility Poor reproducibility of separation patterns and peak response has been a concern for capillary electrophoresis in general and protein CE in particular since its early development in the 1980s. It was widely appreciated that protein-wall interactions were largely responsible for variable EOF and the resulting variability in migration times and peak areas. As a consequence, the development of means to control or eliminate wall interactions continues to be a major research effort in CEo Pioneering efforts by Hjerten, Regnier, Schomburg, El Rassi,
Englehardt, and others lead to the development of separation chemistries and capillary coatings which provided successful protein separations in many applications. At this writing, over a dozen coated capillaries are commercially available. However, there is clearly room for improvement as commercial coated capillaries still do not provide satisfactory lifetime under harsh analysis conditions, e.g., high pH. It is hoped that improved bonding conditions and coating structures as well as improvement in the quality of the fused silica will provide more stable capillaries with reproducible performance. An alternative strategy is the use of rapid in situ coating methods with which a user can replace lost coating periodically, recovering lost performance and greatly extending capillary lifetime at minimal cost.
