title: Characterization of lysozyme adsorption in cellulosic chromatographic materials using small-angle neutron scattering.
abstract: Measurements of the nanoscale structure of chromatographic adsorbents and the associated distribution of sorbed protein within the media can facilitate improvements in such media. We demonstrate a new technique for this purpose using small-angle neutron scattering (SANS) to characterize the nano- to microscale structure of the chromatographic media and sorbed protein under conditions relevant for preparative chromatographic separations. The adsorption of lysozyme on cellulosic S HyperCel™ (Pall Corporation), a strong cation exchanger, was investigated by SANS. The scattering spectrum is reduced to three contributions arising from (1) the chromatographic medium, (2) discrete protein molecules, and (3) the distribution of sorbed protein within the medium. These contributions are quantified for a range of protein loadings. The total concentration of protein in the chromatographic media can be quantified from the SANS spectrum and the protein is observed to retain its tertiary structure upon adsorption, within the resolution of the method. Further analysis of the SANS spectra shows that protein adsorption is uniform in the media. These measurement techniques provide new and valuable nanoscale information about protein sorption in chromatographic media.
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of Delaware, Newark, DE 19716, USA.
authors.forename: Stijn H S
authors.initials: SH
authors.lastname: Koshari
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of Delaware, Newark, DE 19716, USA.
authors.forename: Norman J
authors.initials: NJ
authors.lastname: Wagner
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of Delaware, Newark, DE 19716, USA. Electronic address: lenhoff@udel.edu.
authors.forename: Abraham M
authors.initials: AM
authors.lastname: Lenhoff
mesh_terms: D000327:Adsorption; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D002845:Chromatography; D009113:Muramidase / Q000032:analysis / Q000737:chemistry*; D033363:Neutron Diffraction; D009502:Neutrons; D011506:Proteins / Q000032:analysis; D053838:Scattering, Small Angle
chemical_list: D011506:Proteins; D009113:Muramidase
keywords: Architecture and performance determination; Cellulosic chromatographic media; Protein adsorption measurement; Protein chromatography; Small-angle neutron scattering