Multiple myeloma: Detection of free monoclonal light chains by modified immunofixation electrophoresis with antisera against free light chains

Abstract

Introduction Neoplastic monoclonal gammopathies are frequently associated with production of excess free monoclonal light chains. A sensitive method for detecting free monoclonal light chains in serum could provide a marker for residual/minimal residual disease and as an adjunct to serum protein electrophoresis to serve as a screening method for monoclonal gammopathies. Methods Conventional serum immunofixation electrophoresis was modified by applying undiluted serum samples, and staining for serum free light chains with antisera specific to free light chains. Washing/blotting of gels was enhanced to remove residual proteins that did not bind to the antibodies including intact monoclonal immunoglobulins. Results from this modified immunofixation electrophoresis were compared with results from commercially available MASS-FIX/MALDI assay. Results Monoclonal free kappa light chains could be detected to a concentration of about 1.78 mg/L and monoclonal free lambda light chains to a concentration of about 1.15 mg/L without the need for special equipment. Comparison of these thresholds with parallel results from a commercially available MASS-FIX/MALDI assay revealed the modified electrophoretic method to be more sensitive in the context of free monoclonal light chains. Conclusions Modified serum immunofixation electrophoresis has been shown to detect low levels of monoclonal free light chains at a sensitivity suitable for the method to be used in detecting minimal residual disease, and potentially in a screening assay for monoclonal gammopathies. The disparity in the results with commercially available MASS-FIX/MALDI assay is postulated to be due to limited repertoire of reactivity of nanobodies of camelid origin.

The potential role of mass spectrometry for the identification and monitoring of patients with plasma cell disorders: Where are we now and which questions remain unanswered?

“…This need for accurate background information was highlighted in the study by Wilhite et al where they reported that a sample tested by MASS-FIX at the Mayo clinic was reported to contain elotuzumab when the patient was not being treated with this tmAb. 63 The identification of a light chain with a m/z consistent with a tmAb does not definitively confirm that presence of the tmAb and should be reviewed in the context of the medical and treatment history.…”

“…There are case examples where MASS-FIX outperformed Freelite 61 and other cases where Freelite provided greater sensitivity. 62,63 In light of the variable sensitivity for the detection of monoclonal FLC compared to the serum FLC assays, it is important that serum FLC and BJP assessments continue to be used alongside the MALDI-TOF MS assays that do not include reagents specifically designed for the detection of monoclonal FLC.…”

“…Suitability for large-scale adoption would also need to be explored as the additional laboratory technician time was approximately 1 h per gel. 63 Disease assessment based on FLC assessments in patients with renal impairment can be particularly challenging. The kappa to lambda ratios observed in patients with renal impairment are increased and a wider renal reference range has to be used for the interpretation of serum FLC ratios in this patient cohort.…”

“…The assays do not specifically measure the monoclonal FLC and rely on skewing the FLC ratio to detect monoclonal FLC, so results are more difficult to interpret in the context of renal impairment due to the reduced clearance rate Intact light chain MALDI-TOF MS 145 63 Currently only available in the US Next-generation flow cytometry 450 18 Requires a bone marrow biopsy procedure to obtain a suitable sample for analysis Samples need to be processed within 24-48 h of collection Next-generation sequencing 1950 86 Requires a bone marrow biopsy procedure to obtain a suitable sample for analysis PET-CT 870 95 Able to monitor intra-and extramedullary disease…”

“…The much lower polyclonal total light chain background signal at the tails compared to the apex of the m/z range may explain the variable sensitivity of the MASS‐FIX assay for the detection of low‐level monoclonal FLC when compared to Freelite. There are case examples where MASS‐FIX outperformed Freelite 61 and other cases where Freelite provided greater sensitivity 62,63 . In light of the variable sensitivity for the detection of monoclonal FLC compared to the serum FLC assays, it is important that serum FLC and BJP assessments continue to be used alongside the MALDI‐TOF MS assays that do not include reagents specifically designed for the detection of monoclonal FLC.…”