Performance validation of mass spectrometry for detecting urinary Bence Jones protein and evaluation of its clinical application in multiple myeloma and amyloidosis

OBJECTIVES: Traditional urine immunofixation electrophoresis (uIFE) is cumbersome, has limited sensitivity, and is subjectively interpreted. This is especially problematic in renal impairment where accumulation of polyclonal free light chains may pseudonormalize the serum free light chain (sFLC) ratio, compromising its reliability for disease monitoring. This study aimed to validate two MALDI-TOF mass spectrometry (MS)-based methods for urinary M-protein detection and to evaluate their clinical utility, particularly in addressing this limitation of sFLC ratio interpretation. METHODS: In 325 patients with multiple myeloma or amyloidosis, we compared two conventional uIFE methods (uIFE and uIFE-FLC) with two novel MALDI-TOF MS methods (iMS-IP Assay-urine and iMS-FLC Assay-urine). Patients were divided into a uIFE-positive cohort (164 cases) and a uIFE-negative cohort (161 cases). We evaluated the agreement between the novel methods and serological evidence (serum immunofixation electrophoresis [IFE] and sFLC), as well as their performance in challenging scenarios including renal impairment. RESULTS: The iMS-IP Assay-urine showed excellent agreement with uIFE (94.46 %, κ=0.877) and detected additional M-protein-positive cases that correlated with serum findings. The iMS-FLC Assay-urine showed moderate agreement with uIFE-FLC (80 %, κ=0.615) and detected significantly more positive cases, with support from serum assays. Notably, in six seronegative patients with renal impairment, iMS-FLC Assay-urine detected monoclonal free light chains in all cases, whereas uIFE-FLC detected only three. Both mass spectrometry methods also provided N-glycosylation information. CONCLUSIONS: The iMS-IP and iMS-FLC urine assays are sensitive, rapid, and objective, outperforming conventional uIFE. They offer critical complementary information, particularly in seronegative patients with renal impairment, supporting their potential to replace traditional urine immunofixation electrophoresis.