Reducing heterophilic antibody interference in immunoassays using single-chain antibodies

• Published in: Analytical biochemistry Vol. 419; no. 2; pp. 333 - 335
• Main Authors: Baird, Cheryl L., Tan, Ruimin, Fischer, Christopher J., Victry, Kristin D., Zangar, Richard C., Rodland, Karin D.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 15.12.2011
• Subjects: 60 APPLIED LIFE SCIENCES; Animals; ANTIBODIES; Antibodies, Heterophile - immunology; biomarkers; blood serum; diagnostic techniques; ELISA; ENZYME IMMUNOASSAY; enzyme-linked immunosorbent assay; FUNCTIONALS; HAAA; HAMA; Humans; Immunoassay - methods; Immunoassay interference; Immunoglobulin G - blood; Mice; microarray technology; PATIENTS; POLYPEPTIDES; Protein microarray; scFv; scFv, immunoassay, microarray, ELISA; Single-Chain Antibodies - blood; Single-Chain Antibodies - immunology; TARGETS; HAAA; Protein microarray; HAMA; scFv; Immunoassay interference; ELISA
• ISSN: 0003-2697; 1096-0309
• DOI: 10.1016/j.ab.2011.08.025

Sandwich enzyme-linked immunosorbent assay (ELISA) microarrays can simultaneously quantify the levels of multiple diagnostic targets in a biological sample. However, as with traditional ELISA diagnostics, endogenous antibodies in patient sera can cause interference. We demonstrate here that reducing the diagnostic capture antibody to its minimal functional unit (i.e., a single-chain antibody fragment [scFv]) is an effective strategy for reducing assay interference. Our finding illustrates a source of error introduced by the reliance on immunoglobulin-based capture reagents in sandwich immunoassays with human serum samples. We demonstrate that scFvs can be used in such assays to improve reliability by reducing heterophilic antibody interference, thereby improving biomarker analysis and validation.