Validation of an LC-Q-TOF “HiRes Mass-fix” assay for improved sensitivity and specificity of M-protein detection

Abstract Objectives To validate a blood-based liquid chromatography quadrupole time-of-flight mass spectrometry (MS) method “HiRes Mass-Fix” as a fit-for-purpose, patient-specific method for detecting low level M-proteins in patients who have reached negativity by MALDI-TOF MS “Mass-Fix”. Methods Analytical validation included precision, accuracy, analytical measurement interval, limit of detection, specificity, ion suppression, carryover, and extracted-sample stability. Clinical performance was assessed by comparing 174 patients with bone marrow (BM) measurable residual disease (MRD) by next-generation flow cytometry (NGF) to both Mass-Fix and HiRes Mass-Fix. Results HiRes Mass-Fix demonstrated acceptable precision, accuracy, linearity, specificity, and stability for its intended use, with an average limit of detection of 3.8 mg/L. HiRes Mass-Fix detected M-proteins in 41 of 58 Mass-Fix negative samples. This raised blood MRD positivity from 62 to 89 % against a BM NGF positivity rate of 51 %. Blood-BM agreement fell from 71 % (kappa 0.43) by Mass-Fix to 58 % (kappa 0.15) by HiRes Mass-Fix, driven by blood-positive/BM-negative discordance. IgG was overrepresented among discordant cases (68 vs. 45 %, p=0.003). Conclusions HiRes Mass-Fix was successfully validated and demonstrated to increase M-protein detection in samples negative by Mass-Fix. IgG recycling may explain the blood-positive/BM-negative discordance, and serial measurements will be needed to distinguish residual M-protein from ongoing production. Blood-based M-protein measurement is a less invasive, lower-cost alternative to bone marrow biopsy for ongoing patient monitoring.