Quantitation of serum monoclonal proteins: Relationship between agarose gel electrophoresis and immunonephelometry

BACKGROUND: Previous comparisons of monoclonal protein quantification identified a bias between serum protein electrophoresis (SPEP) and immunonephelometry (NEPH). METHODS: We reviewed data from 2845 patients in whom a single sample provided a γ fraction M-spike by SPEP, a heavy chain isotype by immunofixation electrophoresis (IFE), and an Ig quantification by NEPH. We examined the relationship between SPEP and NEPH. Selected sera with high monoclonal protein concentrations were diluted and reassessed. RESULTS: For all isotypes, the relationship between SPEP and NEPH was best fitted with cubic curves. We determined the concentrations of each isotype that fitted a linear relationship. IgA had the best correspondence (slope 0.92, 95% CI 0.87-1.02), whereas IgM demonstrated a systematic bias of higher values by NEPH (slope 1.80, 95% CI 1.68 -1.92). IgG demonstrated a nonlinear relationship between SPEP and NEPH, with a linear region <19.2 g/L having a slope of 0.83 (95% CI 0.79-0.89) and a second linear region having a slope of 1.47 (95% CI 1.39 -1.53) at higher concentrations. Dilutions of high-concentration IgG monoclonal proteins were linear by NEPH and nonlinear by SPEP. CONCLUSIONS: There are systematic differences in the quantification of monoclonal IgM and IgG by SPEP and NEPH. The bias in IgM is from NEPH overestimation. The nonlinearity of SPEP at high monoclonal IgG concentrations may obscure changes in plasma cell populations. Clinicians should be made aware of the biases and nonlinearity in these tests to make proper conclusions regarding treatment response.