Fig E1.

Detection of functionally active antibodies in culture supernatants by using flow cytometry. A, Schematic depicting the principle for antibody detection in cell-culture supernatants by using a classical sandwich ELISA (ELISA) and our novel cytofluorometry-based assay (flow cytometry). B and C, Standard curves showing ranges of linearity (dotted lines) for ELISA (Fig E1, B) and for flow cytometry (Fig E1, C). D, Comparison between IgE concentrations calculated with ELISA and flow cytometry. Error bars represent SEMs of 3 independent experiments. E, Screening of clones transfected with pVITRO1-vector or UCOE-vector system graph representing secreted anti-CSPG4 IgE detected with flow cytometry. Clones secreting between 2 and 4 μg/mL anti-CSPG4 IgE were considered medium-expressing clones, and those that produced greater than 4 μg/mL were considered high-expressing clones. The right panel summarizes absolute numbers and percentages of different antibody expression levels.