Challenges of measuring monoclonal proteins in serum

Abstract

Abstract:The measurement of monoclonal protein (M-protein) is vital for stratifying risk and following individuals with a variety of monoclonal gammopathies. Direct measurement of the M-protein spike by electrophoresis and immunochemical measurements of specific isotypes or free light chains pairs has provided useful information about the quantity of M-protein. Nonetheless, both traditional electrophoresis and immunochemical methods give poor quantification with M-proteins smaller than 10 g/L (1 g/dL) when in the presence of polyclonal immunoglobulins that co-migrate with the M-protein. In addition, measurements by electrophoresis of M-proteins migrating in the β-and α-regions are contaminated by normal serum proteins in those regions. The most precise electrophoretic method to date for quantification involves exclusion of the polyclonal immunoglobulins by using the tangent skimming method on electropherograms, which provides a 10-fold improvement in precision. So far, however, tangent measurements are limited to γ migrating M-proteins. Another way to improve M-protein measurements is the use of capillary electrophoresis (CE). With CE, one can employ immunosubtraction to select a region of interest in the β region thereby excluding much of the normal proteins from the M-protein measurement. Recent development of an immunochemical method distinguishing heavy/light chain pairs (separately measuring IgGK and IgGL, IgAK and IgAL, and IgMK and IgML) provides measurements that could exclude polyclonal contaminants of the same heavy chain with the uninvolved light chain type. Yet, even heavy/light results contain an immeasurable quantity of polyclonal heavy/ light chains of the involved isotype. Finally, use of liquid chromatography-tandem mass spectrometry (LC-MS/MS) looms on the horizon as a means to provide more consistent and sensitive measurements of M-proteins.

Protein characterization using electrophoresis and immunofixation; a case‐based review of dogs and cats

“…A total protein measurement is needed to convert protein fraction percentages into a gravimetric protein measurement . Often laboratories will use the biuret, Lowery, or other automated dye‐binding protein assays already in place on their chemistry analyzer to determine the total protein in serum samples submitted for electrophoresis .…”

“…With this technique, the sample is incubated with an antibody against the target molecule(s) prior to electrophoresis. When electrophoresed, the target molecule‐labeling immunoglobulin complexes have altered migration, causing a relative void in the electrophoretogram when compared with the electrophoretogram of the sample without labeling . In effect, the target protein has been subtracted from the electrophoretogram.…”

Paraproteínas: claridad en la nebulosa

“…Dado que existen pocas causas de hipogammaglobulinemia, (inmunodeficiencias, posterior a fármacos como rituximab), este hallazgo en la EFP sumado a una clínica compatible, debe hacer sospechar en una GM como la primera causa. Es decir, un paciente que presenta hipogammaglobulinemia y alta sospecha clínica, obliga a buscar la paraproteína con técnicas más sensibles 4 .…”

Protein electrophoresis and serum free light chains in the diagnosis and monitoring of plasma cell disorders: laboratory testing and current controversies

“…In this next group of papers and case reports on PCD, Keren and Schroeder begin by reviewing the electrophoretic and immunochemical methods that have been used over the years to quantify M-proteins in serum [11]. They explore the limitations of past and current methods and describe new methods to improve the accuracy of measurement of low-concentration M-proteins and quantify isotype-specific immunoglobulin classes, and liquid chromatography-tandem mass spectrometry to obtain more analytically sensitive measurements of residual M-protein.…”

“…As discussed in Section 2 [11], one way to improve the accuracy of measurement of low-concentration M-proteins includes use of immunoassays that quantify isotype-specific immunoglobulin classes. In their "Point" paper, Evans et al [31] describe the advantages of using IgA-κ and IgA-λ heavy/light chain (HLC) assays as a more accurate method for quantification of monoclonal IgA proteins that overlap normal proteins in the β-region on electrophoresis.…”