Accuracy of serum IgM and IgA monoclonal protein measurements by densitometry

We previously reported that proper dilution of sera that contain IgG monoclonal proteins (M-proteins) is necessary for accurate quantitation of serum albumin and M-protein concentrations separated by serum protein electrophoresis (SPE) using the Beckman PARAGON agarose electrophoresis system. We now report the significance of pre-electrophoretic serum dilution for M-protein quantitation of sera from patients with IgA and IgM monoclonal gammopathy. We measured M-proteins by SPE in 82 serum samples from 29 patients with IgA and 72 samples from 23 patients with IgM monodonal gammopathy. The serum M-protein concentrations (mean +/- SD) at 1:5, 1:10, and 1:20 dilutions (v/v) for all samples of both types were 49.7 +/- 12.9, 49.1 +/- 13.1, and 47.8 +/- 13.0 g/L, respectively. Thirty-two (20.8%) of 154 sera showed varying degrees of increase in M-protein concentrations with serum dilutions higher than 1:5; only 8 (5.2%) showed an increase 3 SDs. By SPE, the M-protein concentration (mean +/- SD) of these 8 sera at 1:5, 1:10, and 1:20 dilutions were 52.6 +/- 7.8, 57.1 +/- 7.2, and 57.6 +/- 7.1 g/L, respectively; the albumin concentrations (mean +/- SD) were 41.4 +/- 4.4, 37.9 +/- 3.8, and 37.1 +/- 2.9 g/L, respectively. The corresponding albumin concentration (mean +/- SD) was 36.8 +/- 3.7 g/L, assayed by the bromcresol green dye-binding method. These 8 samples were obtained from 3 patients, 2 with IgM kappa and 1 with IgA lambda monoclonal gammopathy. On the electrophoresis membranes, the M-protein bands of these 8 samples were narrow, thin, and dense; upon scanning, they appeared taller and thinner than the corresponding albumin bands. The samples of this subset contained relatively high concentrations of M-protein and total serum protein. We conclude that a pre-electrophoretic dilution of 1:5 (v/v) is adequate for most sera with IgA or IgM M-proteins. However, 1:10 or 1:20 dilution is occasionally required for a subset of sera with IgA or IgM M-proteins that show an unusually thin, narrow, and dense M-protein band and have high total serum protein content.