DOI: 10.5061/dryad.8tk26
title: Data from: Assessment of intrathecal free light chain synthesis: comparison of different quantitative methods with the detection of oligoclonal free light chains by isoelectric focusing and affinity-mediated immunoblotting

description: Objectives: We aimed to compare various methods for free light chain (fLC)
 quantitation in cerebrospinal fluid (CSF) and serum and to determine
 whether quantitative CSF measurements could reliably predict intrathecal
 fLC synthesis. In addition, we wished to determine the relationship
 between free kappa and free lambda light chain concentrations in CSF and
 serum in various disease groups. Methods: We analysed 166 paired CSF and
 serum samples by at least one of the following methods: turbidimetry
 (Freelite™, SPAPLUS), nephelometry (N Latex FLC™, BN ProSpec), and two
 different (commercially available and in-house developed) sandwich ELISAs.
 The results were compared with oligoclonal fLC detected by
 affinity-mediated immunoblotting after isoelectric focusing. Results:
 Although the correlations between quantitative methods were good, both
 proportional and systematic differences were discerned. However, no major
 differences were observed in the prediction of positive oligoclonal fLC
 test. Surprisingly, CSF free kappa/free lambda light chain ratios were
 lower than those in serum in about 75% of samples with negative
 oligoclonal fLC test. In about a half of patients with multiple sclerosis
 and clinically isolated syndrome, profoundly increased free kappa/free
 lambda light chain ratios were found in the CSF. Conclusions: Our results
 show that using appropriate method-specific cut-offs, different methods of
 CSF fLC quantitation can be used for the prediction of intrathecal fLC
 synthesis. The reason for unexpectedly low free kappa/free lambda light
 chain ratios in normal CSFs remains to be elucidated. Whereas CSF free
 kappa light chain concentration is increased in most patients with
 multiple sclerosis and clinically isolated syndrome, CSF free lambda light
 chain values show large interindividual variability in these patients and
 should be investigated further for possible immunopathological and
 prognostic significance.
description: Sample Blot
 1Sample Blot
 2Sample Blot
 3Raw data in Excel
 Tableflc
 102015 essential data for DRYAD.xlsAnalytical
 characteristics of the in house ELISA methodsAnalytical
 characteristics of the in house ELISA methods – Table Legend CoH – fKLC
 Freelite high control; CoL – fKLC Freelite low control; CSF1, CSF2, CSF3 –
 pooled CSF samples; S1, S2, S3 – pooled serum samples; 1/10, 1/100, 1/250
 – pooled CSF dilutions. Pooled sera as well as Freelite controls were
 diluted 1/1000. SD – standard deviation; CV % – coefficient of variation.
 Flebogamma 40 mg/L was added to test maximal theoretical cross-reaction
 with light chains bound in the IgG. There is, however, no information
 available whether intravenous immunoglobulin preparations may contain
 traces of free light chains. Please note that in Method D, sample S1 was
 out of the calibration range of the fLLC assay. Estimated bias is only
 provided for method E since there is no internationally recognized
 standard for fLC; hence, Freelite controls were presumed to give the
 indicated concentrations only in conjunction with Freelite calibrators.
 Recovery assays: One of the pooled CSFs and sera was spiked with 10%
 (vol/vol) of either Bethyl purified monoclonal fLC or Freelite standards
 to obtain the theoretical concentration indicated (i.e., B16 corresponds
 to the final concentration of 16 ug/L in the sample). The expected
 concentration was then calculated as the final theoretical concentration +
 0.9 * concentration measured in the non-spiked sample within the same
 analytical run. Recovery was calculated as 100 * (measured concentration –
 expected concentration)/expected concentration.FLC SPA
 preanalytics11 CSF samples were
 divided in two aliquots; one was frozen to -30°C immediately upon receipt
 (A), the second was stored at +2 to +8°C for exactly 7 days and frozen
 thereafter. All samples were then analysed for free light chains within
 the same analytical run using the Freelite assay on SPA PLUS
 analyser.fKLC
 preanalytics11 CSF samples were
 divided into two aliquots and frozen either immediately (A) or after 7
 days at +2°C to +8°C (B). Only marginal, albeit statistically significant,
 decrease of fKLC concentrations has been found in (B).fLLC
 preanalytics11 CSF samples were
 divided into two aliquots and frozen either immediately (A) or after 7
 days at +2°C to +8°C (B). No changes of fLLC concentrations have been
 observed in (B).