Validation of an Immunoprecipitation, Digestion, and Immunoaffinity LC/MS/MS Assay for Human beta-NGF Biomarker and Implementation in Support of Clinical Trials

摘要

A quantitative immunoaffinity LC/MS/MS assay was developed and validated for total human beta-NGF. Unlike typical immunoassays, only a single reagent antibody is required for enrichment, reducing assay interference of the therapeutic antibody. Advanced assay validation was performed using a 7.0 to 450 pg/mL calibration range. Nanoelectrospray stability over 35 hours with a 10 minute cycle time. The inter-run accuracy ((percent)RE) ranged from -15.2percent to 16.3percent across all validation samples. Inter-run assay precision ((percent)CV) ranged from 7.0percent to 21.2percent. The assay has proven to be sensitive (sub-picomolar) and robust (2percent run failure rate).