Surface plasmon resonance spectroscopy-based high-throughput screening of ligands for use in affinity and displacement chromatography.
We describe an approach that uses surface plasmon resonance (SPR) spectroscopy and self-assembled monolayers (SAMs) for the high-throughput screening of ligands for use in displacement and affinity chromatographic processes. We identified a set of commercially available organic amines and allowed them to react with SAMs presenting interchain carboxylic anhydride groups; the resulting surfaces presented ligands of interest in a background of carboxylic acid groups. We used SPR spectroscopy to determine the extent of adsorption of two model proteinslysozyme and cytochrome conto these "multimodal" surfaces and to select promising "affinity" ligands for further characterization. The attachment of selected ligands to UltraLink Biosupport resulted in beads with a significantly greater affinity for lysozyme than for cytochrome c that would be suitable for use in affinity chromatographic processes. Furthermore, we also used the screens to design "affinity displacers"small molecules that selectively retain lysozyme on chromatographic resins, while displacing cytochrome c. The combination of SPR spectroscopy and SAMs represents a powerful technique for identifying novel ligands that enable the purification of complex protein mixtures.
Vutukuru
Srinavya
S
Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, New York 12180, USA.
Kane
Ravi S
RS
D000327:Adsorption; D000588:Amines / Q000737:chemistry; D000818:Animals; D002627:Chemistry, Physical / Q000379:methods; D002645:Chickens; D002845:Chromatography / Q000379:methods*; D045304:Cytochromes c / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D006736:Horses; D008024:Ligands; D009113:Muramidase / Q000737:chemistry*; D012492:Salts / Q000737:chemistry; D020349:Surface Plasmon Resonance / Q000379:methods*
D000588:Amines; D008024:Ligands; D012492:Salts; D045304:Cytochromes c; C421864:hen egg lysozyme; D009113:Muramidase