High-sensitivity Quantitative Analysis of a Monoclonal Antibody in Human Serum Using Q Exactive Selected Ion Monitoring NanoLC/MS

摘要

A research mAb was quantified in human serum by nanoLC-Q Exactive MS with both SIM and targeted MS/MS methods without using any isotope-labeled internal standards. (1) Using targeted SIM, the LLOQ of 2 ng/mL in human serum for the antibody was achieved with good precision (CV < 10%) and good accuracy (Diff < 25%) (n=3). The mass accuracy is less than 2 ppm for all present isotopes of targeted peptide. (2) The SIM quantitation has similar performance with respect to the sensitivity (2 ng/mL LLOQ), dynamic range (2-6250 ng/mL) and linearity (R2 > 0.996) compared to the quantitative results generated from a triple quadruple LC-MS system (Data not shown). (3) Targeted MS/MS method provided an alternative way and improved selectivity to quantify peptides in a more complex matrix. One or more fragment ions might be employed for highly sensitive quantitation. The peptide sequence could be confirmed from the MS/MS spectrum and data can be re-analyzed without further sample analysis. In short, these methods provide high specificity, ability to multiplex while requiring short assay development time. The SIM nanoLC-MS strategy can be a highly sensitive and robust method for therapeutic mAb quantitative analysis, and has a great potential for use in support of pharmacokinetic studies of therapeutic proteins in early and late stages of drug development.