title: Method for studying immunoglobulin G binding on hydrophobic surfaces.
abstract: We used a reactant adsorptive membrane bioreactor separator (or RAMBS) system to examine hydrophobic interaction based binding of human immunoglobulin G (HIgG) on synthetic microporous membranes possessing tunable hydrophobicity. Membrane bound HIgG on being pulsed with papain resulted in Fab being obtained in the flowthrough with Fc remaining bound to the membrane. On the other hand, when membrane bound HIgG was pulsed with pepsin, Fc subfragments were obtained in the flowthrough with F(ab')(2) remaining bound to the membrane. These product profiles suggest that HIgG bound to the membrane through its middle region. Enzyme linked immunoadsorbent assay (ELISA), sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and mass spectrometric analysis of eluate samples obtained from the RAMBS experiments provided evidence that the binding of HIgG took place primarily through the segment consisting of the hinge and C(H)2 domain of Fc. The experimental approach described in this paper could potentially be more widely applicable for studying protein interactions with membrane and surfaces in general.
authors.affiliation: Department of Chemical Engineering, McMaster University, 1280 Main Street West, Hamilton, Ontario, Canada L8S 4L7.
authors.forename: Deqiang
authors.initials: D
authors.lastname: Yu
authors.forename: Raja
authors.initials: R
authors.lastname: Ghosh
mesh_terms: D019149:Bioreactors; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D007074:Immunoglobulin G / Q000737:chemistry* / Q000378:metabolism; D013058:Mass Spectrometry; D008567:Membranes, Artificial; D008962:Models, Theoretical; D010206:Papain / Q000378:metabolism; D010434:Pepsin A / Q000378:metabolism; D011145:Polyvinyls / Q000737:chemistry; D011485:Protein Binding; D005465:Fluorocarbon Polymers
chemical_list: D007074:Immunoglobulin G; D008567:Membranes, Artificial; D010206:Papain; D010434:Pepsin A; D011145:Polyvinyls; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers