A1. Affinity ranking of antibodies by BIAcore.
A2. Affinity determination of clone 2 antibody in different concentration (nM)
A GenScript produced Rabbit anti-ID antibody generally has high affinity at picomolar.
B. Blocking assay
B Competitive ELISA with different concentration of drug target could set as an optional step to distinguish the type of anti-ID antibodies. Ab1 and Ab2, which could bind to the antibody drug when competed with drug target, are non-blocking anti-ID antibodies, while Ab3 and Ab4 are antigen-blocking antibodies.
|Capture mAb, 2μg/ml||Concentration of Antibody Drug (ng/ml)||Detection mAb,1μg/ml|
C. Pairing antibody selection.
C Sandwich ELISA could be performed to select anti-ID pair which could largely increase the success rate of immunoassay development. As it was shown in the table, mAb2 is able to pair with other four clones (mAb1, mAb3, mAb4, mAb5).
D Purified anti-idiotype polyclonal antibodies showed specific binding to antibody drug, not to isotype control. The cross rate with isotype control is less than 10%.