title: Label-free Fab and Fc affinity/avidity profiling of the antibody complex half-life for polyclonal and monoclonal efficacy screening.
abstract: A unified approach to affinity screening for Fab and Fc interactions of an antibody for its antigen and FcγR receptor has been developed. An antigen array is used for the Fab affinity and cross-reactivity screening and protein A/G proxy is the FcγR receptor. The affinities are derived using a simple 1:1 binding model with a consistent error analysis. The association and dissociation kinetics are measured over optimised times for accurate determination. The Fab/Fc affinities are derived for ten antibodies: mAb-actin (mouse), pAb-BSA (sheep), pAb-collagen V (rabbit), pAb-CRP (goat), mAb-F1 (mouse), mAbs (mouse) 7.3, 12.3, 29.3, 36.3 and 46.3 raised against LcrV in Yersinia pestis. The rate of the dissociation of antigen-antibody complexes relates directly to their immunological function as does the Fc-FcγR complex and a new half-life plot has been defined with a Fab/Fc half-life range of 17-470 min. The upper half-life value points to surface avidity. Two antibodies that are protective as an immunotherapy define a Fab half-life >250 min and an Fc half-life >50 min as characteristics of ideal interactions which can form the basis of an antibody screen for immunotherapy.
authors.affiliation: Biosciences, College of Life and Environmental Sciences, University of Exeter, Stocker Road, Exeter, EX4 4QD, UK.
authors.forename: Thomas
authors.initials: T
authors.lastname: Read
authors.affiliation: Biosciences, College of Life and Environmental Sciences, University of Exeter, Stocker Road, Exeter, EX4 4QD, UK.
authors.forename: Rouslan V
authors.initials: RV
authors.lastname: Olkhov
authors.affiliation: Biomedical Sciences Department, Defence Science and Technology Laboratory, Porton Down, Salisbury, Wiltshire, SP4 0JQ, UK.
authors.forename: E Diane
authors.initials: ED
authors.lastname: Williamson
authors.affiliation: Biosciences, College of Life and Environmental Sciences, University of Exeter, Stocker Road, Exeter, EX4 4QD, UK. andrew.m.shaw@exeter.ac.uk.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Shaw
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000493:pharmacokinetics; D000915:Antibody Affinity; D003429:Cross Reactions; D006207:Half-Life; D007140:Immunoglobulin Fab Fragments / Q000276:immunology*; D007141:Immunoglobulin Fc Fragments / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments
keywords: Bioanalytical methods; Immunoassays/ELISA; Optical sensors