Analysis of biomolecular interactions using affinity microcolumns: A review

Authors

Xiwei Zheng, University of Nebraska–LincolnFollow
Zhao Li, University of Nebraska–LincolnFollow
Sandya Beeram, University of Nebraska–LincolnFollow
Maria Podariu, University of Nebraska–Lincoln
Ryan Matsuda, University of Nebraska–LincolnFollow
Erika L. Pfaunmiller, University of Nebraska–LincolnFollow
Christopher J. White II, University of Nebraska–Lincoln
NaTasha Carter, University of Nebraska–Lincoln
David S. Hage, University of Nebraska - LincolnFollow

Date of this Version

1-2014

Citation

Published in Journal of Chromatography B (2014); doi: 10.1016/j.jchromb.2014.01.026

Comments

Copyright © 2014 Elsevier B.V. Used by permission.

Abstract

Affinity chromatography has become an important tool for characterizing biomolecular interactions. The use of affinity microcolumns, which contain immobilized binding agents and have volumes in the mid-to-low microliter range, has received particular attention in recent years. Potential advantages of affinity microcolumns include the many analysis and detection formats that can be used with these columns, as well as the need for only small amounts of supports and immobilized binding agents. This review examines how affinity microcolumns have been used to examine biomolecular interactions. Both capillary-based microcolumns and short microcolumns are considered. The use of affinity microcolumns with zonal elution and frontal analysis methods are discussed. The techniques of peak decay analysis, ultrafast affinity extraction, split-peak analysis, and band-broadening studies are also explored. The principles of these methods are examined and various applications are provided to illustrate the use of these methods with affinity microcolumns. It is shown how these techniques can be utilized to provide information on the binding strength and kinetics of an interaction, as well as on the number and types of binding sites. It is further demonstrated how information on competition or displacement effects can be obtained by these methods.