Bedřich Friedecký

Interlaboratory study of free monoclonal immunoglobulin light chain quantification

Abstract Background: Quantification of monoclonal immunoglobulin free light chains (FLCs) in serum is used increasingly in clinical practice for the diagnosis, prognostic assessment, and treatment monitoring of monoclonal gammopathies. It is used as an adjunct to standard serum protein electrophoresis and immunofixation. However, methods for FLC quantification need further standardization and validation. Methods: The Czech Myeloma Group and the Czech Society of Clinical Biochemistry have initiated an interlaboratory study where six laboratories collaborating with the primary myeloma treatment centres measured FLC concentrations in 12 serum samples from patients with monoclonal gammopathies. Results: Repeatability of the measurements in five laboratories was calculated based on differences between the results of duplicate measurements. We found that repeatability depended more on the laboratory than on the device used for measurement. Conclusions: The study revealed several weak points in the methodology, including the need for a uniform sample dilution procedure. Interlaboratory reproducibility was comparable with values achieved in the NEQAS programme. Because the κ/λ ratio cannot be measured with high precision, κ and λ FLC concentrations should be used where possible. Due to its impact on the clinical management of patients with gammopathy, FLC quantification needs to become a part of the regular quality control cycle in myeloma centres.

Diagnostic Kits Derived from Standard Method "DGKC 94" as a Potential Tool for Improvement of Analytical Standardization and Clinical Utility of Alkaline Phosphatase

Sir, The harmonization of methods used for the measurements of catalytic concentration (enzymatic activity) of alkaline phosphatase is still far from ideal. In the past years, several solutions have been proposed as the way out from this well-known dilemma. In 1983, the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) proposed a recommended method, using 4-nitrophenol phosphate as a substrate and 2-amino-4-methylpropanol as a buffer at an incubation temperature of 30 °C (1). However – as just recently demonstrated – commercial methods based on this recommendation produce significant differences in results when patient’s sera are measured (2). These differences are usually not recognized by schemes of external quality assessment which use lyophilized control material. On the other hand, the inter-laboratory comparison of results from the same patient may confuse clinical interpretation. The reasons for such discrepancies are in differences in reagent formulation of the commercial kits, including the variations of the sample/reagent ratio and concentrations of reagents, and the presence or absence of additives such as magnesium, zinc and ethylenediaminetetraacetate (EDTA), and the other substances potentially able to influence the activity of alkaline phosphatase. For instance use of diethanolamie buffer instead of 2-amino-4-methylpropanol can produce up to two-fold higher values (2). An alternative procedure for the measurement of the alkaline phosphatase was proposed in 1981, using N-methyl-D-glucamine buffer. This substance can be prepared in crystalline-pure form, which can help to reduce the potential problem of impurities (3). The method was recommended for standardization in Italy (4) and later successfully tested in the USA (5). Finally, in 1992 it was proposed by the German Society of Clinical Chemistry (DGKC) as the “DGKC 94” standard method (6). This non-patented method is carried out at 37 °C and it is popular due to its robustness. In order to check the analytical performance of its commercial adaptations, we evaluated the following three kits with respect to their precision, accuracy and reproducibility. (A)“Alkalische Phosphatase (Standardmethode 94) – BM 1662961”, Boehringer Mannheim (Roche), Germany; (B)“Alkalische Phosphatase (DGKC new), Ecoline 25”, Merck, Germany; (C)“Alkalická fosfatáza BLT 220”, Lachema, Czech Republic.