The heavy and light chains of the 5A6 antibody and the CD81 large extracellular loop were co-expressed in Expi293F cells: - 600 mL of Expi293F cells maintained in Expi293 expression medium were grown to a density of 2.8 × 106 cells/mL and then transiently transfected with heavy chain 5A6, light chain 5A6, and CD81 LEL DNA (0.48 mg total DNA) and FectoPro transfection reagent (Polyplus) at a 1:1 DNA/FectoPro ratio. - 20 hr after transfection, the cells were fed 5 mM Valproic acid sodium salt and 5.5 mL of 45% D-(+)-Glucose solution. - Transfected cells were cultured for 7 days to produce protein and then the medium was collected and separated from the cells by centrifugation at 4000 g for 15 min at 4°C. The cultured medium was loaded onto protein A resin. Expression and purification of Ab5, Ab10, Ab21, and 5A6 for use in Flow Cytometry: - Expi293F cells maintained in Expi293 expression medium were grown to cell density of 2.8 × 106 cells/mL and then transiently transfected. - For each antibody, plasmids encoding the heavy and light chain were transfected at a 1:2 molar ratio (0.8 mg total DNA/liter cells) with FectoPro transfection reagent (Polyplus) at 1:1 DNA/FectoPro ratio. - 20–24 hr after transfection, the cells were fed with 5 mM Valproic acid sodium salt and 45% D-(+)-Glucose solution. - 4–7 days after transfection, the medium was collected and separated from the cells by centrifugation at 4000 g for 15 min at 4°C. Cultured medium was loaded onto protein A resin.