Development of an anti-drug antibody assay to detect anti-drug antibodies to protein and PEG in a PEGylated molecule

Background: PEGylation technology is one of long-acting delivery (LAD) platforms used to increase half-life of protein therapeutics. However, PEGylation of anti-Factor D Fab (PEG-aFD) poses challenges for detecting anti-drug antibody (ADA) to both Fab and polyethylene glycol (PEG) portions. Results: Although the bridging ELISA using traditional assay diluent containing Tween 20 is good for detecting ADA to Fab, it failed to detect ADA to PEG. Instead of only reducing Tween 20 in assay diluent, using a proprietary commercial buffer PEG50-1 as assay diluent successfully enabled the detection of ADA to both Fab and PEG with fit-for-purpose sensitivity and drug tolerance. Conclusion: Identification of appropriate assay diluent is critical for detection of ADA to both Fab and PEG in a PEGylated molecule.