Quality assurance for serum free light chain analysis

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28.1. Introduction

Figure 28.1 QA003 Distribution 115, February 2011. A. SPE gel accompanying report. Lanes 2-5: Normal human serum (NHS); Lanes 6-9: QA003 serum. B and C. sIFE gels accompanying report. Fκ: Free kappa antisera. This sample contained monoclonal κ FLCs in the absence of monoclonal intact immunoglobulin.

Figure 28.2 QA003 Distribution 115, February 2011. Frequency distribution of reported κ sFLC (Freelite) results. Red histogram: all participants; blue histogram: BNII and ProSpec results. The result from the reporting laboratory is indicated by an arrow.

Figure 28.3 QA003 Distribution 115, February 2011. Combined sFLC (Freelite) results represented on a dot plot. The result from the reporting laboratory is indicated by an arrow.

Figure 28.4 NEQAS Distribution 94. κ Freelite CV strongly influenced by a single outlier (indicated by *) which was not trimmed from the analysis.

Figure 28.5 NEQAS Distribution 95. κ Freelite results show bimodal distribution due to antigen excess. Some participants reported κ concentrations from initial sample dilution (falsely low due to antigen excess), others reported true value.

It is essential that all laboratories participate in external quality assurance (QA) schemes when performing monoclonal protein analysis. There are several national and international QA schemes for assessing serum free light chains (sFLC). Particular care should be taken when reporting sFLC results: in the USA mg/dL units are commonly used, while elsewhere mg/L is the norm. Individual QA scheme results that are 10- or 100-fold different from the majority view may be attributed to mathematical errors in calculating the sample concentration.

28.2. The Binding Site scheme

The Binding Site IMMPROVE™ serum paraprotein QA scheme (QA003) was the first scheme to include sFLC analysis. To date, there have been more than fifty distributions over 10 years, and around 300 laboratories worldwide currently participate in the scheme.

Four serum samples are issued per year and results can be returned via the IMMPROVE website. Methodologies assessed include serum protein electrophoresis (SPE), serum immunofixation electrophoresis (sIFE) and Freelite™ sFLC assays. Laboratories are able to return quantitative results for IgG, IgA, IgM, β₂-microglobulin, κ sFLC, λ sFLC and κ/λ ratio. A comprehensive report is returned to each laboratory. Freelite™ results are categorised according to nephelometric/turbidimetric instrument type to allow user group comparisons to be made. The report also includes examples of electrophoresis gels and an expert opinion regarding the sample and results. Electrophoresis and sFLC results from a typical QA003 report (Distribution 115, February 2011) are shown in Figures 28.1 to 28.3.

28.3. United Kingdom National External Quality Assessment Service scheme

The United Kingdom National External Quality Assessment Service (UK NEQAS) for Immunology provides a Monoclonal Protein Identification scheme that has incorporated sFLC analysis since 2005. Over 350 UK and international laboratories currently participate in the scheme. There are 6 distributions per year comprising unmatched serum and urine samples.

In its current format, the sFLC scheme does not exclude extreme outliers from the analysis, and overall %coefficient of variation (CV) results may be strongly influenced by an individual participant's discrepant results (Figure 28.4). Past distributions have also highlighted the need for all participants to be aware of sFLC assay technical issues, including antigen excess. The κ sFLC results returned for NEQAS Distribution 095 were bimodal. For the BN™II instrument which does not include automatic antigen excess checks, two peaks were observed at 143 mg/L and 1001 mg/L (Figure 28.5). It is likely that such discrepant results correspond to those participants who did and those who did not screen for sFLC antigen excess. The phenomenon of antigen excess is further discussed in Chapter 4, Section 4.2 I and the recommended sample dilution protocol is outlined in the Freelite™ product insert.

28.4. College of American Pathologists scheme

The College of American Pathologists (CAP) has recently introduced a proficiency testing scheme for sFLCs. There are 3 distributions per year, and currently over 150 participants are enrolled in the scheme. In contrast to The Binding Site and NEQAS schemes described above, the CAP sFLC scheme does not include proficiency testing for serum electrophoresis, which is tested separately.

28.5. Randox International Quality Assessment scheme

The Randox International Quality Assessment scheme (RIQAS) for FLCs, initiated in 2007, was the first clinical external quality control scheme to gain United Kingdom Accreditation Service (UKAS) accreditation. Twelve serum samples are distributed every 6 months. Over 90 samples have been distributed to date, and there are currently around 40 participants in the scheme.

28.6. German Institute for Standardisation scheme

The German Institute for Standardisation's Gesellschaft zur Förderung der Qualitätssicherung in Medizinischen Laboratorien scheme (abbreviated to INSTAND e.V.) provides a monoclonal protein scheme with over 760 participating laboratories reporting IFE. Since 2007, sFLCs have been added and there are currently over 70 users. Two plasma samples are distributed 4 times per year.