Figure 1. Generation of mAbs specific for rhAMHR2-ED.
A panel of approximately 300 hybridoma supernatants were generated and screened for specificity against rhAMHR2-ED and the 4D12 parental hybridoma was selected for subcloning by limiting dilution. (A) Subcloning produced three sub-clones, 4D12C6, 4D12C7, and 4D12G1 each of which expressed the IgG1/κ-chain isotype and showed antigen specificity (B) by competitive ELISA and (C) by flow cytometry binding to OVCAR8 cells. For flow cytometry, positive control staining of OVCAR8 cells was performed using a commercially available anti-AMHR2-ED mAb (Abcam), whereas IgG1 isotype antibodies with irrelevant specificities were used as negative controls. In all cases, error bars indicate ± SD and the results shown are representative of three experiments yielding similar results.