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David F Keren, Commentary, Clinical Chemistry, Volume 55, Issue 3, 1 March 2009, Pages 580–581, https://doi.org/10.1373/clinchem.2008.118596
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These cases remind us that IgM monoclonal proteins (M-proteins) have earned a reputation as saboteurs of many laboratory assays owing to their self-aggregation, aggregation of latex particles, binding to analytes and reagents, and cryoprecipitation. One additional point is that some IgM M-proteins also play havoc with their own detection and measurement by electrophoretic techniques.
Immunoelectrophoresis, an older technique still used by a dozen or so laboratories, employs a long gel-diffusion step during which large IgM molecules move through agarose more slowly than the sleeker IgG, resulting in a phenomenon known as the umbrella effect. This effect results in false negatives when the IgM M-protein light chain is obscured by the polyclonal light chains of IgG (1). The solution employed for this problem is to break the disulfide bonds with 2-mercaptoethanol. Immunoelectrophoresis has been largely replaced by immunofixation. One of the many advantages of immunofixation is that the reagent antisera are placed directly on the gel, so a long diffusion phase is not required and the umbrella effect is virtually eliminated. Another problem with IgM measurement has emerged recently with capillary zone electrophoresis. With this technique, IgM M-protein measurement occasionally yields markedly lower quantities than those obtained with nephelometry or densitometric scan of serum protein electrophoresis on agarose gels (2). As with the umbrella effect in immunoelectrophoresis, capillary zone electrophoresis measurement of the IgM M-protein is improved by pretreatment with 2-mercaptoethanol, suggesting that factors relating to size or self-aggregation may interfere with the passage of M-proteins through the narrow capillary. In my laboratory we recommend that the first time an IgM M-protein is identified by capillary zone electrophoresis, it should be subjected to agarose gel electrophoresis to compare the quantity of the M-protein. If there is a disparity, we recommend following the M-protein by densitometric scans using agarose gel electrophoresis or by nephelometry for measuring total IgM concentration. There remains an important role for keen scrutiny of data by laboratorians to prevent spurious results from affecting clinical decisions.
