The effect of paraprotein polymerisation on quantitation by capillary zone electrophoresis and Hevylite®.

OBJECTIVES

Up to 3% of patients with monoclonal gammopathies have multiple serum paraproteins. This article investigates whether multiple isotype-matched paraproteins, as seen on capillary zone electrophoresis, are truly biclonal.

METHODS

Serum samples containing multiple isotype-matched paraproteins were treated with the reducing agent dithiothreitol, and capillary zone electrophoresis was performed pre- and post-treatment. Band resolution and effect of resolution on quantitation of paraprotein burden were assessed. The Hevylite turbidimetric assay was also evaluated for ability to quantify such paraproteins.

RESULTS

Among patients with biclonal isotype-matched paraproteins, 23/24 (96%) IgA paraproteins resolved into a single band following treatment with dithiothreitol compared with only 1/12 (8%) IgG paraproteins. Daratumumab therapy accounted for the second band in 5/9 non-resolving IgGκ paraproteins. Where initially quantified as a single IgA 'complex' (multiple bands in close proximity), the single postdithiothreitol band averaged 2.8 g/L less (<0.001), likely due to inclusion of lower amounts of underlying serum proteins (y = 0.97x-2.03, R=0.993). Quantitating IgA biclonal isotype matched ( = 58) using the Hevylite assay gave higher results ( = 0.002) than capillary zone electrophoresis (y = 1.48x-7.13, R=0.959). In contrast, single IgA paraprotein results ( = 48) did not differ between the two methods ( = 0.466; y = 1.24x-2.74, R=0.898), suggesting that polymerisation enhances Hevylite quantitation.

CONCLUSIONS

These results suggest that disulphide-mediated polymerisation of IgA paraproteins is more common than true biclonal gammopathy and support dithiothreitol treatment of samples with isotype-matched IgA bands before quantifying by capillary zone electrophoresis. The Hevylite assay should be utilized with caution where polymerisation is likely. Where IgGκ biclonal isotype-matched paraproteins appear on capillary zone electrophoresis, daratumumab therapy should be considered.