Serum Free Light Chain and Drift: Calibrator Adjustment Needed?

Serum free light chains (sFLC), which include kappa (KFLC) and lambda (LFLC), and their ratio (K/L ratio) are used for diagnosis and monitoring of plasma cell dyscrasias. Previously, The Binding Site Freelite® assay established reference intervals for KFLC, LFLC, and K/L ratio, in healthy donor (1) and K/L ratio for chronic kidney disease (CKD) populations (2). We read with great interest the article “Kappa Free Light Chain Drift Prompts the Need for a New Upper Limit of Normal Free Light Chain Ratio to Avoid an Epidemic of Kappa Light Chain Monoclonal Gammopathy of Undermined Significance” by Rozenova et al. who described drift of the Kappa Freelite assay from The Binding Site (3). Assay drift for Freelite has been reported before (4), suggesting the need to verify and/or re-establish these intervals in clinical practice, while some manufacturers have not recommended re-establishing K/L ratio reference intervals for the renal-impaired population (5). We recently evaluated healthy and renal insufficiency population reference intervals for sFLC on the Optilite platform. Two hundred apparently healthy serum samples were procured from BioIVT as a normal sample set with ages representing those that are encountered in clinical practice for suspected plasma cell dyscrasia and 60 samples from Precision BioSciences from a cohort known to have high creatinine values. All 260 serum samples were measured using the Freelite assay on the Optilite. Creatinine was measured on the Abbott Alinity and estimated glomerular filtration rate (eGFR) was calculated using the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) 2021 equation. This cohort was split into 2 groups: eGFR ≥60 (n = 181) and eGFR ≤59 (n = 79). In the ≥60 eGFR cohort (n = 181), the sFLC medians were compared to the Optilite product insert medians, which are referenced to the 2002 Katzmann paper (1). The median recovered for KFLC was 22.5 mg/L, which is approximately 308% above the claimed median of 7.3 mg/L. For LFLC, the median recovered was 17.9 mg/L, which is approximately 145% above the claimed median of 12.4 mg/L. The median recovered for K/L ratio was 1.27, approximately 211% above the claimed median of 0.60. To assess if this large percent drift using commercially available normal serum samples relates to clinical patient data, approximately 105 000 data points each of KFLC and LFLC from our institution using Optilite since 2017 were graphed on the KFLC vs LFLC plot in log form, with normal ranges for KFLC, LFLC, and K/L ratio superimposed in black lines, as well as the medians in red lines (Fig. 1). Recalibrated results were also graphed by applying division factors of 3.1 to KFLC (to account for 310% drift) and 1.45 to LFLC (to account for 145% drift). The Institutional Review Board at the University Health Network in Toronto, Ontario, Canada deemed the study exempt from review. Over 100 000 patient sFLC results from Freelite on Optilite are plotted in log scale. The published reference intervals for KFLC, LFLC, and K/L ratio are depicted by the black solid lines and the medians found in our eGFR ≥60 cohort are shown by the solid red lines. The recalibrated dataset (factor of 3.1 for KFLC, 1.45 for LFLC) is shown on the right with the claimed medians in solid red lines; all units are in mg/L. The medians found in the ≥60 eGFR n = 181 cohort (KFLC: 22.5 mg/L, LFLC: 17.9 mg/L, and K/L ratio: 1.27) coincide with the center of a high density of clinical patient data points that represent an ellipse, which also has roughly the same width of the 0.26 to 1.65 K/L ratio and appears shifted to the right. When recalibrating the Freelite assay using the factors from the median drift, the medians from the 2002 Katzmann study can be recovered for normal serum samples for Freelite on Optilite, as well as align with the center of the high density of clinical patient data points. It is possible that after recalibration is performed the need for renal K/L ratio reference intervals may not be necessary for Freelite as the median ratio of 0.60 is fairly centered about the 0.26 to 1.65 range. Since recalibrating an assay by 300% is a sizable shift from the current state and the ratio of drift between KFLC and LFLC is a factor of approximately 2, adjusting the KFLC assay down 200% will also center Freelite to the 0.60 median, with the concomitant result that the medians for both KFLC and LFLC would still be about 150% above normal median claims. In conclusion, rather than redefine K/L ratio ranges (6) and lose the historical significance and clinical validation of the established 0.26 to 1.65 K/L ratio, an alternate solution is to redefine the Freelite lambda and/or kappa calibrator values such that normal ranges for normal eGFR groups (≥60) can be recovered with another benefit of possibly not needing redefined K/L ratio reference ranges for abnormal eGFR groups (≤59). Nonstandard Abbreviations: KFLC, kappa free light chains; LFLC lambda free light chains; K/L, kappa/lambda ratio; eGFR, estimated glomerular filtration rate. Author Contributions:The corresponding author takes full responsibility that all authors on this publication have met the following required criteria of eligibility for authorship: (a) significant contributions to the conception and design, acquisition of data, or analysis and interpretation of data; (b) drafting or revising the article for intellectual content; (c) final approval of the published article; and (d) agreement to be accountable for all aspects of the article thus ensuring that questions related to the accuracy or integrity of any part of the article are appropriately investigated and resolved. Nobody who qualifies for authorship has been omitted from the list. Mark Griffiths (Conceptualization-Equal, Data curation-Equal, Formal analysis-Equal, Visualization-Equal, Writing—original draft-Equal, Writing—review & editing-Equal), Randal Schneider (Conceptualization-Equal, Funding acquisition-Equal, Investigation-Equal, Project administration-Equal, Resources-Equal, Validation-Equal, Writing—review & editing-Equal), and Vathany Kulasingam (Conceptualization-Equal, Data curation-Equal, Formal analysis-Equal, Investigation-Equal, Methodology-Equal, Project administration-Equal, Supervision-Equal, Validation-Equal, Writing—review & editing-Equal) Authors’ Disclosures or Potential Conflicts of Interest:Upon manuscript submission, all authors completed the author disclosure form. Research Funding: Abbott Diagnostics provided the reagents and funding to complete this study. Disclosures: V. Kulasingam serves as a consultant for Abbott and is an Associate Editor for The Journal of Applied Laboratory Medicine, Association for Diagnostics & Laboratory Medicine (ADLM). R. Schneider is an employee of Abbott, receives stock or stock options from Abbott, and sits on the Policy & External Affairs Core Committee for ADLM.

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