ABSTRACT
Separation technology has been central to the elucidation of protein structure and function. Both chrolnatography and electrophoresis have been used for decades to isolate and characterize proteins and their cOlnponents. Open column and low pressure chronlatography were essential for the purification of sufficient amounts for protein for structural analysis, and the preparation of high-resolution ion exchange resins enabled developlnent of the alnino acid analyzer which paved the way for analysis of protein composition. The introduction of protein-compatible ion exchange and size exclusion supports allowed high-performance liquid chromatography to be used for obtaining highly pure proteins, while high-efficiency reversed phase columns proved invaluable for high-resolution peptide mapping and purification, and for identification of the PTH amino acids generated by automated Edman sequence analysis. Gel electrophoresis evolved in parallel with chromatography, providing an inexpensive method for separating complex protein mixtures. The development of the Laemmli system for separating SDS-protein complexes on polyacrylamide gels proved to be such a powerful technique that it is used on a daily basis in virtually every protein chemistry laboratory in the world. Isoelectric focusing provided an alternative separation technique for separation of proteins based on their isoelectric points, and the combination of IEF and SDS-PAGE by O'Farrell resulted in a two-dimensional separation technique which has the power to resolve thousands of proteins on a single 2-D gel. The development of blotting techniques to transfer separated proteins from the gel to a
suitable support for other analyses such as immunoassay or sequencing greatly simplified many experiments in protein chemistry.
