1 of 33

Journal Presentation

Dr. Md. Raiq Raihan Chowdhury

Resident

Department of Haematology

BSMMU

2 of 33

Occurrence of Double Monoclonal Bands on Protein Electrophoresis: An Unusual Finding

Vishrut K Srinivasan, Priyanka Bhagat, Frainey Bansal, Seema Chhabra

​

Indian J Hematol Blood Transfus

(June 2016) 32 (Suppl 1):S184–S188

3 of 33

Background

​

  • Monoclonal gammopathy represents presence of Monoclonal protein (M-protein) in serum and/or urine

​

​

​

  • A result of overproduction of immunoglobulin by a clone of abnormally proliferating B lymphocytes or plasma cells

4 of 33

Background (contd.)

  • Monoclonal gammopathy can be found in:
    • Monoclonal gammopathy of undetermined significance
    • Multiple Myeloma
    • Waldenstorm’s Macroglobulinemia
    • Primary amyloidosis
    • Cryoglobulinemia
    • Lymphoproliferative disorders
    • Solitary plasmacytoma
    • Infections i.e, Hepatitis C
    • POEMS syndrome

​

5 of 33

Background (contd.)

​

  • Immunofixation electrophoresis (IFE) is recommended for definitive identification of monoclonal proteins and characterization of heavy chain (HC) and light chain (LC) being secreted

​

​

6 of 33

Introduction

  • In a certain subset of patients with monoclonal gammopathies, more than one M-protein can be found in serum and/or urine.

​

  • This category of gammopathies are called ‘double gammopathies’.

​

  • The term ‘biclonal gammopathy’ has been used interchangeably in literature, in lieu of ‘double gammopathy’.

7 of 33

Introduction

  • Double gammopathy may present with two separate distinct M band/peak on SPEP or a single M band on SPEP which further resolves into two separate bands on IFE.

​

  • The double gammopathies are further subdivided depending on the type of HC and LC detected.

8 of 33

Introduction

  • IFE in these cases can reveal either different HC isotypes with same LC isotype or same HC isotype with dissimilar LC isotypes

​

​

  • The subset of cases with different HC isotypes with different LC isotypes has been distinctively referred to as “true biclonal gammopathy”

9 of 33

Conditions associated with double gammopathy

Double gammopathies are known to be associated with

  • multiple myeloma
  • certain lymphoproliferative disorders
  • ‘double/biclonal gammopathy of undetermined significance’ (DGUS/BGUS), similar to MGUS

10 of 33

Origin of Double Gammopathy

​

  • Either detected at initial presentation or during the course of the disease after initial diagnosis has been established.

​

​

  • Can result from proliferation of ‘‘two or more distinct plasma cell clones’’ producing more than one monoclonal protein.

​

11 of 33

Origin of Double Gammopathy

​

  • Alternatively, ‘‘clonally related’’ plasma cells which have undergone class switch recombination at a later stage in disease pathogenesis can also give rise to double gammopathies.

​

​

  • During disease progression or following the treatment, the level of the ‘double’ M-proteins may rise or fall in concordant or discordant manner

​

12 of 33

Incidence of Double Gammopathies

​

  • The incidence of double gammopathies among various cohorts of gammopathies in reported literature is 2–6 %

​

​

  • IgG–IgA has been reported as the most common HC isotype combination by Kyle et al.

​

13 of 33

Incidence of Double Gammopathies

​

  • Riddell et al. observed IgG–IgM to be the most common HC isotype combination.

​

​

  • Certain other studies have found occurrence of IgG–IgG as the most common combination

14 of 33

Incidence of Double Gammopathies

​

  • The differences between these case series may be the result of
    • difference in population distributions,
    • racial differences,
    • differences in sensitivity of detection method used, or
    • selection bias during case selection for inclusion in the series.

​

​

  • Among the light chains, kappa light chain was most commonly detected

15 of 33

  • True biclonal gammopathies may result either from neoplastic proliferation of two different ‘‘clonally unrelated’’ malignant plasma cell lines or from the existence of a single clone of B-lymphoid cells, which subsequently diversifies into two independent clones after a process of antigenic selection

​

  • Accordingly, the clonal proteins being secreted always have a ‘‘different variable region and differ in both light chain and heavy chain class’’

16 of 33

  • provides conclusive evidence that the two M-bands seen are actually the product of two separate clones and represent a ‘true’ biclonal gammopathy

​

  • The detection of a single LC isotype in majority of double gammopathies, as mentioned earlier, is suggestive of a single clonal origin of M-proteins in these cases

17 of 33

18 of 33

19 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

  • In certain situations, two distinct bands on SPEP yield only a single band in one of the HC (IgG/IgM/IgA) lanes along with two bands in a single light chain lane i.e. either kappa or lambda on IFE

​

​

  • One of the two bands seen in light chain lane is due to the presence of excess free LCs which migrate faster compared to LCs associated with Ig heavy chains.

​

20 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

​

​

​

  • This could be due to asynchronous production of the components of the Ig molecules, leading to synthesis of an intact monoclonal Ig plus excess monoclonal free LCs

21 of 33

22 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

  • Similarly, resolution of the subtypes causing two bands may also be complicated especially in case of IgA gammopathies.

​

​

​

  • Monoclonal IgA molecules have a tendency to dimerize, and the resulting dimer often has a different mobility than the monomer parent molecule, thereby, yielding two bands on SPEP in some cases of IgA monoclonal gammopathies.

​

23 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

  • In these circumstances, IFE will yield two bands in IgA lane at different positions along with corresponding 2 bands in one of the LC lanes i.e. IgA kappa IgA kappa

​

​

  • Pre-treatment of sera with 2-mercaptoethanol depolymerizes the multimers and helps in better distinction of double gammopathies

24 of 33

25 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

  • the pentameric IgM can break down into 7 s subunits, which show up on electrophoresis as one or more ‘extra’ monoclonal bands.

​

  • These multiple bands are most likely due to various molecular forms of a monoclonal protein derived from a single plasma cell clone

26 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

​

  • In certain cases of IgA myeloma, there may not be any corresponding band in LC lane at the level of IgA band in the HC lane.

​

​

​

  • This finding can be reported as ‘‘IgA with no apparent or discernible light chain attached’’.

​

27 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

​

  • This occurs due to structural properties of IgA molecule that hinders the reaction between LC epitopes and the antibodies against them

​

  • This may also occur due to prozone phenomenon where there is excess of antigen or low antibody concentration

28 of 33

Does a Double Band Always Signify Biclonal/ Double Gammopathy?

  • Presence of additional band in beta-gamma region on SPEP can sometimes interfere with analysis and maybe misinterpreted as a monoclonal band/peak.

​

  • This band is caused by presence of fibrinogen which can occur due to insufficient clotting of sample.

​

  • However, this ‘fibrinogen’ band disappears on IFE as it does not bind to any specific antisera used.

​

  • Thus, in such situations, IFE is helpful in reaching a diagnostic conclusion.

29 of 33

30 of 33

Utility of Nephelometry

  • The quantitative FLC assay by nephelometry is more sensitive than electrophoretic techniques for detection of serum FLC.

​

​

  • In addition to LCs, HCs can also be quantified with the help of nephelometric methods.

​

​

  • This may aid in better identification of separate Ig classes in cases of biclonal/double gammopathies.

31 of 33

Implications

  • Clinically, there is no difference between double/biclonal gammopathy and monoclonal gammopathy in terms of outcome for the patient

​

​

  • However, recognition of this entity not only increases diagnostic accuracy and credibility, but also helps in assessing treatment response during follow-up of patients.

32 of 33

Implications

  • Measurement of both clonal proteins, at diagnosis and during follow-up, helps in identifying synchronicity of response to both clones.

​

  • The difference in response of different clones can aid in driving further treatment decisions in such cases.

  • Additionally, the recognition of the different monoclonal components can serve as a tool for further deciphering disease pathogenesis and for defining a distinct subset of monoclonal gammopathies.

33 of 33

Thank You