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Targeted Antibody Blocking by a Dual-Functional Conjugate of Antigenic Peptide and Fc-III Mimetics DCAF
Published on: September 17, 2019
Matrix interference from Fc-Fc interactions in immunoassays for detecting human IgG4 therapeutics
Michael A Partridge1, Elif Kabuloglu Karayusuf1, Gangadhar Dhulipala1
1Regeneron Pharmaceuticals, Inc., 777 Old Saw Mill River Road, Tarrytown, NY 10591, USA.
Background:
An assay measuring an IgG4 biotherapeutic in human serum used a drug-specific monoclonal antibody (mAb) capture reagent and an antihuman IgG4 mAb as detection reagent. However, serum IgG4 binding to the capture mAb via Fc-interactions was detected by the anti-IgG4 mAb, causing high background.
Results:
Two approaches were developed to minimize background; incorporating a mild acid sample preparation step or using the Fab of the capture antibody. Either strategy improved signal:noise dramatically, increasing assay sensitivity >20-fold. Biophysical analyses of antibody domains indicated that noncovalent Fc oligomers could inhibit the background.
Conclusion:
Matrix interference from human IgG4 binding to the capture mAb was reduced with a Fab fragment of the drug-specific capture antibody or by incorporating a mild acid sample treatment into the assay.
