The different molecules were stably expressed, as previously described [30 ]. CHO-S, HEK293 or YB2/0 cells were stably transfected with the appropriate linearized expression vectors. Ch12G4, h12G4 and 3C23K antibodies were produced in YB2/0 cell using EMS (Invitrogen), 5% Ultra-low IgG fetal calf serum (FCS) (PAA) and 0.5g/l G418 for 5 to 7 days. 3C23K-CHO-S was produced in CHO-S cells using ProCHO4 (Lonza), 4mM glutamine and 1g/l G418 for 7 days.
MAbs were purified from culture supernatants by affinity chromatography using protein A sepharose (GE-Healthcare). The levels of aggregates and endotoxins were determined by gel filtration on Superdex HR/200 (GE-Healthcare) and by LAL testing, respectively. Antibody quality and purity were monitored by SDS-PAGE and Coomassie staining. In addition, the glycosylation patterns and core fucose percentage of each purified antibody were determined by high performance capillary electrophoresis laser induced fluorescence (HPCE-Lif) [55 (link)].
MAbs were purified from culture supernatants by affinity chromatography using protein A sepharose (GE-Healthcare). The levels of aggregates and endotoxins were determined by gel filtration on Superdex HR/200 (GE-Healthcare) and by LAL testing, respectively. Antibody quality and purity were monitored by SDS-PAGE and Coomassie staining. In addition, the glycosylation patterns and core fucose percentage of each purified antibody were determined by high performance capillary electrophoresis laser induced fluorescence (HPCE-Lif) [55 (link)].
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