New Methods and Applications for the Chromatography of Biological Macromolecules
Chromatography is widely used for both the analytical-scale and process-scale separation of biological macromolecules. The most common way to perform this application of chromatography involves using gradients in fluid composition where the gradient is produced by mixing conducted external to the column. However, rather than using these mechanically produced gradients, another option is to employ gradients that are chemically produced inside the column by exploiting the titration of ionogenic functional groups on the column packing and the adsorption behavior of the species in the liquid phase forming the gradient. Although much less widely practiced due to its complexity, this latter technique, termed chromatofocusing, has a number of useful properties, such as the ability to produce focusing effects that enhance the resolution, the ability to retain the biological activity of an eluted species, and the ability to perform highly reproducible separations at very small scales, such as in a microfluidic device.
This seminar will begin with an overview of focusing methods in the general field of separation science, and the fundamental principles underlying chromatofocusing in particular will be described. Results will then be presented from recent work on novel chromatofocusing techniques applied to biopolymers with computer-aided methods used to optimize the separation. Applications of these methods will be described which combine electrostatic interactions with hydrophobic interactions to improve the resolution of proteins, and which combine affinity interactions with focusing effects to minimize the co-elution of impurities with the affinity target. Applications will also be described for performing quality control in monoclonal antibody production and for performing multidimensional chromatography for proteomic analysis.
Douglas Frey is currently a professor in the Department of Chemical, Biochemical, and Environmental Engineering at the University of Maryland Baltimore County (UMBC) in Baltimore, Maryland. He obtained his B.S. degree from Stanford University and his M.S. and Ph.D. degrees from the University of California, Berkeley, all in chemical engineering. Dr. Frey has conducted both fundamental and applied research on chromatography and related topics for nearly 30 years, first as an assistant professor and then as an associate professor at Yale University where he collaborated with Csaba Horváth, and more recently as a full professor at UMBC. Dr. Frey's work encompasses both experimental and theoretical research as well as the development of computer-aided-design software for chromatography, including software that has been sold commercially in the biotechnology industry. Dr. Frey was the chair of the Department of Chemical, Biochemical, and Environmental Engineering at UMBC from 1996-2001 and the program director for the Separation Processes Program at the National Science Foundation from 2001-3 and again from 2005-6. He is currently on the editorial board of the journal Biotechnology and Applied Biochemistry. He is also works with several companies and governmental agencies as a consultant and collaborator, including W.R. Grace, Genzyme, Genentech, and the U.S. Food and Drug Administration.