title: A biotin-drug extraction and acid dissociation (BEAD) procedure to eliminate matrix and drug interference in a protein complex anti-drug antibody (ADA) isotype specific assay.
abstract: Monitoring anti-drug antibody (ADA) responses in patients receiving protein therapeutics treatment is an important safety assessment for regulatory agencies, drug manufacturers, clinicians and patients. Recombinant human IGF-1/IGFBP-3 (rhIGF-1/rhIGFBP-3) is a 1:1 formulation of naturally occurring protein complex. The individual IGF-1 and IGFBP-3 proteins have multiple binding partners in serum matrix with high binding affinity to each other, which presents challenges in ADA assay development. We have developed a biotin-drug extraction with acid dissociation (BEAD) procedure followed by an electrochemiluminescence (ECL) direct assay to overcome matrix and drug interference. The method utilizes two step acid dissociation and excess biotin-drug to extract total ADA, which are further captured by soluble biotin-drug and detected in an ECL semi-homogeneous direct assay format. The pre-treatment method effectively eliminates interference by serum matrix and free drug, and enhances assay sensitivity. The assays passed acceptance criteria for all validation parameters, and have been used for clinical sample Ab testing. This method principle exemplifies a new approach for anti-isotype ADA assays, and could be an effective strategy for neutralizing antibody (NAb), pharmacokinetic (PK) and biomarker analysis in need of overcoming interference factors.
authors.affiliation: Bioanalytical and Biomarker Development, Shire, Lexington, MA 02421, United States. Electronic address: hniu@shire.com.
authors.forename: Hongmei
authors.initials: H
authors.lastname: Niu
authors.affiliation: Bioanalytical and Biomarker Development, Shire, Lexington, MA 02421, United States.
authors.forename: Thomas
authors.initials: T
authors.lastname: Klem
authors.affiliation: Bioanalytical and Biomarker Development, Shire, Lexington, MA 02421, United States.
authors.forename: Jinsong
authors.initials: J
authors.lastname: Yang
authors.affiliation: Bioanalytical and Biomarker Development, Shire, Lexington, MA 02421, United States.
authors.forename: Yongchang
authors.initials: Y
authors.lastname: Qiu
authors.affiliation: Bioanalytical and Biomarker Development, Shire, Lexington, MA 02421, United States. Electronic address: lpan@shire.com.
authors.forename: Luying
authors.initials: L
authors.lastname: Pan
mesh_terms: D000143:Acids; D000906:Antibodies / Q000097:blood*; D057134:Antibodies, Neutralizing / Q000097:blood; D001710:Biotin; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000276:immunology*; D007334:Insulin-Like Growth Factor I / Q000276:immunology*; D011994:Recombinant Proteins / Q000276:immunology; D012680:Sensitivity and Specificity
chemical_list: D000143:Acids; D000906:Antibodies; D057134:Antibodies, Neutralizing; C543394:IGF-I-IGFBP-3 complex; D007074:Immunoglobulin G; D018972:Insulin-Like Growth Factor Binding Protein 3; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D001710:Biotin
keywords: Acid dissociation; Anti-drug antibody (ADA); Electrochemiluminescence (ECL) assay; Immunogenicity; Streptavidin magnetic beads