Abstract

The science, art, and practice of protein purification have been with us for more than a century, yet, in many respects, the field is only now evolving past its adolescent roots. New methods are replacing old methods at such a dizzying pace, that even life-long experts in protein purification cannot keep up. In this article, we present many state-of-the-art protein purification techniques without totally ignoring the past. Our goal is to enable those relatively new to the field of protein purification to choose the best methods to solve their own purification problems. Each method we describe has been used and validated in our own research. We describe these methods, pointing out advantages, disadvantages, and limitations with practical examples rather than with complex, theoretical equations. This paper covers methods of extraction, clarification, batch purification, low pressure column chromatography, HPLC, and electrophoresis as applied to both genetically engineered, recombinant proteins and proteins isolated from natural sources. The relatively new methods of three-phase partitioning, hydrophobic charge induction chromatography, immobilized metal affinity chromatography, and perfusion chromatography are featured.

Keywords: Protein purification, Extraction, Clarification, Chromatography, HPLC, Electrophoresis, Filtration