Research Article
Preexisting Antibodies to an F(ab′)2 Antibody Therapeutic and Novel Method for Immunogenicity Assessment
Figure 2
Preexisting ATAs are directed primarily at the F(ab′)2 hinge, as shown by the reactivity reduction by competition with anti-DLL4 F(ab′)2, Herceptin F(ab′)2, or anti-DLL4 full length antibody in the bridging ELISA format. In the bridging ATA assay using 50 μg/mL competitor (in-well concentration) and a 1/20 sample dilution, full length antibody causes very little signal reduction; however the F(ab′)2 form of the same antibody reduces the signal almost to background. An F(ab′)2 form of Herceptin, which has the same framework as anti-DLL4 but with a different CDR, also reduces the signal, indicating that the signal is due to anti-hinge antibodies.