Comparison of Kappa & Lambda Freelite to Total Kappa & Lambda Immunoassays for the Detection of Monoclonal Gammopathies, Both As Standalone Tests and Alongside Serum Protein Electrophoresis
Authors: Hungria Vania T M , Kampanis Petros , Drayson Mark T , Plant Tim , Crusoe Edvan de Queiroz , Peres Ana Lucia , Cury Priscilla , Sampaio Manuella , Soares Elyara , Harding Stephen 
Published: October 2019

Abstract
Introduction
Monoclonal gammopathies are a disparate group of diseases from benign to malignant which are characterised by the proliferation of a single B cell clone that produces a homogeneous monoclonal immunoglobulin (M-Ig). The method of detection and quantification of the M-Ig will depend upon whether it is an intact immunoglobulin or present as serum free light chain only. Historically serum (SPEP) and urine (UPEP) electrophoresis were considered the gold standard for identifying intact M-Ig and FLC respectively. In 2001 the introduction of the Freelite test changed the diagnostic and monitoring paradigm. The assay is now recommended as a tool to diagnose and monitor patients with B cell disorders. However, the assay is sometimes confused with monospecific immunoassays for measuring total kappa and total lambda. Here we compare kappa & lambda Freelite with total kappa & lambda immunoassays alongside SPEP as tools to identify patients with monoclonal gammopathies.
Materials and Methods
Sera from 102 blood donors (55 males and 47 females, age range 18-67 years) and 103 patients with light chain associated gammopathies (44 males and 59 females, age range 38 to 88 years, 60 kappa / 43 lambda)taken during the course of their treatment were available. The sera was analysed retrospectively with FreeliteTM (The Binding Site Ltd, Birmingham, UK) on a SPAPLUSand Total Kappa &Lambda nephelometricassays (Beckman Coulter, USA) on an Immage.Monoclonality was identified by results falling outside of manufacturers normal ratio ranges (Freelite 0.256-1.65, Total light chain 1.53-3.29). Serum protein electrophoresis was performed and unexpectedly positive or negative results were assessed using immunofixation on the Hydrasys electrophoretic system (Sebia, France).
Results
Monoclonal production was identified in 80/103 light chain associated gammopathiesby Freelite, negative IFE confirmed the absence of monoclonal protein in 22/23 patients with normal FLC kappa/lambda ratios and 1/23 patients had an IgG lambda intact immunoglobulin. SPEP was positive in 30/103 patients, with total kappa/lambda immunoassays detecting monoclonal protein in just 26/103 samples. Freelite was positive in 6/102, SPEP in 2/102 and total kappa/lambda in 8/102 normal blood donor sera. Interestingly, in 1 patient with an abnormal FLC ratio and total kappa/lambda result had a lambda light chain identified using IFE.Comparisons between the performances of Freelite, Freelite + SPEP, Total kappa/lambda and total kappa/lambda + SPEP are shown in table 1).
Conclusion
In keeping with Kyle et al (1999) our study confirms the limitations of total kappa / lambda assays as tools to identify M-Igs. This is the first study looking to apply the recommended algorithm of Freelite + SPEP to the total kappa/lambda assays. The addition of SPEP to total kappa/lambda assays improved the performance to detect abnormalities, but even combined they were neither as sensitive, specific or accurate as the Freelite assay. Given the limitations of the total light chain assays identified in our study, it is important that physicians are aware of which assay is being utilised; one easy method to discriminate would be to look at the normal range of the assay being reported.
Table 1: Comparison of Freelite, Freelite + SPEP, Total kappa/lambda, Total kappa/lambda + SPEP Freelite Freelite + SPEP Total Total + SPEP Sensitivity 77.67 81.55 25.24 43.69 Specificity 94.12 92.16 92.16 91.18 PPV 93.02 91.30 76.47 83.33 NPV 80.67 83.19 54.97 61.59 Accuracy 85.85 86.83 58.54 67.32

Disclosures
No relevant conflicts of interest to declare.