Benchmarking antibody discovery fidelity and reproducibility with an assay-locked residue fidelity index.

Abstract

Reproducibility in antibody discovery is undermined by dropout, paratope degeneracy, and immunogen imprinting. Although guidelines exist, the field lacks a shared, assay-locked metric to benchmark these liabilities across discovery pipelines. We propose the Residue Fidelity Index (RFI), a discovery-stage, within-assay comparative framework that consolidates fidelity liabilities into a single score. Assay-locked means RFI is comparable only within a fully specified assay configuration, uses fixed normalization and weights, and requires co-reporting of component metrics to expose drivers of the composite. RFI is not intended as a universal or optimal standard, but as one implementable example meeting basic benchmarking requirements: defined components, fixed assay context, pre-specified weights, and primary-data disclosure. RFI is reported alongside its components (D, G, I) to visualize reproducibility in parallel with affinity, humanization, and yield. To demonstrate feasibility, we applied RFI in a simulated Epivolve testbed derived from multiple studies, embedding MILKSHAKE (context retention) and Sundae (residue discrimination) validation modules into discovery. Across three campaigns (n = 25 clones), antibodies meeting provisional thresholds (dropout ≤10%, degeneracy ≤0.10, no imprinting) yielded RFI values from 0.610 to 0.982 (mean ± SD 0.922 ± 0.103; I = 1 for clones C04 and C20). These results show how fidelity can be consolidated to prioritize candidates before application-specific cellular validation. Despite limited scope and proprietary immunogens, this work provides a proof of concept for residue-level benchmarking, with per-clone primary values and RFI reported in Supplementary Table S1 for auditability.