Bio-Rad Laboratories Inc, Hercules, Calif, a multinational manufacturer and distributor of life science research products and clinical diagnostics, has produced a technical note that describes the screening, ranking, and epitope mapping of anti-human IL-9 supernatants.
The study described in the technical note, bulletin 5540, demonstrates the screening and ranking of 20 hybridoma supernatants using the company’s SPR biosensor, the ProteOn XPR36 protein interaction array system. Five high-affinity antibodies are identified, and four of these were selected for further purification and epitope mapping.
Two of the antibodies recognize different epitopes on the IL-9 antigen. Epitope mapping and hybridoma ranking were accomplished using the company’s one-shot kinetics approach and the parallel processing capability of the system. The work can be enhanced to include kinetic constant determination of hundreds of supernatants within a few hours.
In the workflow used in the study, five supernatants were screened at a time against five antigen concentrations, and one channel was reserved for referencing. Using this configuration, both the anti-mouse IgG and antibody-containing supernatants were injected into the same set of ligand channels, followed by IL-9 injection into the orthogonal analyte channels.
The company’s one-shot kinetics approach can be expanded to include all six available analyte channels and interspot referencing to conduct supernatant screening. This is accomplished using the ligand channel for immobilization of anti-mouse IgG, the analyte channel for capture of the supernatant clones, and the ligand channels for IL-9 flow.