Monoclonal immunoglobulinS in serum and urine
Each plasma cell clone produces a highly specific monoclonal protein (M protein), which can be used as a tumor marker. No single test can reliably diagnose or monitor all patients.
Detection of monoclonal immunoglobulins in patients with multiple myeloma:
Monoclonal Ig in
serum and urine ----------------- 60%
Monoclonal Ig in serum --------------------------- 20%
Monoclonal Ig in urine ("light-chain myeloma") --- 20%
- Protein electrophoresis (PEP)
- Immunoelectrophoresis (IEP)
- Quantitative immunoglobulins
Immunofixation is a technique which uses electrophoresis and immunodiffusion, and is more sensitive than electrophoresis. Limit of detection: about 10 mg/dL.
PEP should be done in serum, not plasma, because plasma contains fibrinogen, which appears as a discrete band between the beta and the gamma fractions. Serum protein electrophoresis (SPEP) shows the M protein as a peak in the gamma or terminal beta regions. Monoclonal peaks in the alpha or initial beta regions are not due to paraproteins, but to reactant proteins. SPEP can miss the M protein spike if it is very small: limit of detection is about 100 mg/dL.
Examples of proteins separated by SPEP:
- Alpha-1: alpha-1 antitrypsine
- Alpha-2: alpha-2 macroglobulin, haptoglobin, ceruloplasmin
- Beta-1: transferrin
- Beta-2: C3 complement, CRP
- Gamma: immunoglobulins
SPEP is only a screening test; in order to establish the diagnosis of monoclonal gammopathy, an IEP must be done. IEP not only distinguishes polyclonal and monoclonal increases in gammaglobulins, but it also determines the heavy-chain class and the light-chain type of the produced immunoglobulin.
The measurement of the M protein in blood and/or urine is important in assessing the response to the treatment: increasing levels of M protein indicate progression of cancer, while decreasing levels indicate disease regression/remission.
The use of assays to quantitate total kappa and lambda light chains (not to be confused with the free light chains) is not recommended.
Normal serum protein electrophoresis:
Presence of an M spike in the SPEP of a patient with myeloma:
Increase of the M spike over time is consistent with disease progression:
Serum immunofixation of a patient with IgG-lambda myeloma (courtesy of Dr. Michael G. Bayerl - Hematopathology, Penn State Hershey Medical Center):
Urine immunofixation of a patient with myeloma excreting lambda monoclonal proteins (courtesy of Dr. Michael G. Bayerl - Hematopathology, Penn State Hershey Medical Center):
- IgG: about 21 days (except IgG3: 8 days)
- IgA: 6 days
- IgD: 3 days
- IgE: 2 days
- IgM: 5 days
- FLC: about 4 hours (with renal failure: 2-3 days)
Molecular weight of kappa light
chains: 22 kD
Molecular weight of lambda light chains: 45 kD
In SPEP, the protein separation is usually done in agarose gels. In capillary zone electrophoresis (CZE), the proteins are separated in a solution, inside a narrow diameter glass tube.
CZE with immunosubtraction method is more sensitive than the traditional methods for detecting monoclonal proteins. It separates serum proteins after incubation of serum in the presence of antisera specific for heavy chains and light chains. The detection of monoclonal proteins is based on the absence of the removed heavy and light chains when compared to SPEP. The CZE with immunosubtraction method is better than gel electrophoresis because of rapidity, automated report, easy interpretation, and higher sensitivity.
Demonstration of changes in plasma cell
subsets in multiple myeloma.
Haematologica. 2007 Aug;92(8):1135-8.
Ayliffe MJ, Davies FE, de Castro D, Morgan GJ.
Using a double immunofluorescence staining method, these authors demonstrated that bone marrow plasma cells express intact monoclonal immunoglobulins in 74% of case, free light chains in 8% of cases, and a mixture of both cell population in 18% of cases. In a few patients, myeloma cells changed during the course of the disease from producing intact immunoglobulins to cells producing free light chains. The presence of cells expressing free light chains only was associated with worse prognosis.
Monoclonal gammopathy of undetermined significance (MGUS)
consistently precedes multiple myeloma: a prospective study.
Blood. 2009 May 28;113(22):5412-7.
Landgren O, Kyle RA, Pfeiffer RM, Katzmann JA, Caporaso NE, Hayes RB, Dispenzieri A, Kumar S, Clark RJ, Baris D, Hoover R, Rajkumar SV.
Among 77,469 healthy persons enrolled in the nationwide prospective Prostate, Lung, Colorectal, and Ovarian (PLCO) Cancer Screening Trial, 71 persons developed MM. Serum samples were available from 2-9.8 years prior to the diagnosis of MM. MGUS was present in 82% of cases at 8+ years prior to MM diagnosis. Therefore, an asymptomatic MGUS stage preceded MM.
Serum free light chain analysis may miss monoclonal light
chains that urine immunofixation electrophoreses would detect.
Shaheen SP, Levinson SS.
Clin Chim Acta. 2009 Aug;406(1-2):162-6.
Giampaolo Talamo, M.D.