Restricted migration of polyclonal IgG on immunofixation gel electrophoresis in a case of IgG4-related disease.

A 71-year-old male was admitted to the hospital with cough, fatigue and left parotid and left submandibular gland swelling over the previous several months. He had a background history of hypercholesterolaemia on atorvastatin, and hypothyroidism on thyroxine. His blood investigations were nondescript with a <4 mg/L C-reactive protein, and mildly raised erythrocyte sedimentation rate (44 mm/hr). His haematological and biochemical profiles were unremarkable. Fine needle aspirates of the salivary glands revealed reactive lymphoid hyperplasia with no evidence for malignancy. He had a positron emission tomography (PET) scan which showed increased uptake in the parotid glands [standardised uptake values (SUV)max 7.1], the cervical lymph nodes (LN) (SUVmax 6.9), paratracheal LNs (SUVmax 4.1) and mediastinal lymph nodes (SUVmax 4.1). Peripheral blood flow cytometry did not reveal any evidence of B cell clonal disease. Due to concerns of an underlying lymphoproliferative disorder, he had exploratory surgery of his neck, and two enlarged lymph nodes were resected. The samples were sent for histological study and no features were seen to suggest lymphoma, granuloma or metastatic malignancy. However, sections showed reactive lymph nodes with marked capsular and mild parenchymal fibrosis with a background mixed chronic infiltrate including plasma cells, eosinophils and histiocytes. On IgG4/IgG staining, large numbers of plasma cells were present with an IgG4/IgG ratio of >70%. No storiform fibrosis was noted. Serum samples were collected for protein electrophoresis (EPG) and immunofixation electrophoresis (IFE) analysis. This was initially diagnosed by an external laboratory as an IgG-kappa paraprotein. Repeat bloods were performed in our laboratory. The IFE showed a dense focal yet polyclonal migration of IgG in the fast gamma region (Fig. 1A). Subsequently, serum IgG subclasses were measured using SPAPlus turbidimetry which quantified a remarkable IgG4 level of 12.76 g/L (reference range 0.04–0.07). Serum free light chain studies demonstrated a normal kappa:lambda ratio of 1.57 (0.26–1.65), effectively ruling out a monoclonal IgG4 paraprotein. Based on serological and histological findings, the patient was diagnosed with IgG4-related disease (IgG4-RD) and he was treated with prednisolone and methotrexate. Serial EPG/IFE and serum analysis revealed declining IgG4 levels and dissolution of the apparent restricted band (Fig. 1B,C). Six months following treatment, he reported resolution of his cough, fatigue and glandular swelling. To further examine this laboratory phenomena, we examined a series of patients that had similar fast gamma region migrating IgG bands on IFE gel. Five patients were visually identified during a 3-month period in our laboratory. All of these patients were histologically and clinically diagnosed with IgG4-RD and had varying quantities of elevated serum IgG4 level ranging between 3 and 12 g/L. There were no light chain restrictions. We also examined two patients that had IgG4-RD (but IgG4 <2 g/L) that did not have an appreciable fast gamma region-migrating dense band, suggesting that the approximate threshold of visualisation on IFE is an IgG4 level around 2–3 g/L. IgG4-RD is an immune-mediated chronic inflammatory condition of unknown aetiology presenting with mass-forming lesions. These can affect almost any organ and can mimic malignant, infectious and other inflammatory disorders.1Sedhom R. Sedhom D. Strair R. IgG-4 related disease: a mini-review.J Rare Dis Res Treat. 2017; 2: 18-23Crossref Google Scholar IgG4-RD was first described in 2003 in unrelated conditions that shared elevated serum IgG4 levels and common histological findings which included lymphoplasmacytic infiltrate (>40% expressing IgG4 plasma cells), storiform fibrosis and obliterative phlebitis.2Lanzillotta M. Mancuso G. Della-Torre E. Advances in the diagnosis and management of IgG4 related disease.BMJ. 2020; 369: m1067Crossref PubMed Scopus (39) Google Scholar Histological examination of biopsy specimen is the gold standard in confirming IgG4-RD; clinical symptoms and serological findings are needed to confirm diagnosis. Serum IgG4 level is quantified in the laboratory using turbidimetry or nephelometry. About 60–70% of the patients suffering from IgG4-RD have an elevated polyclonal serum IgG4 level.3Jacobs J.F. van der Molen R.G. Keren D.F. Relatively restricted migration of polyclonal IgG4 may mimic a monoclonal gammopathy in IgG4-related disease.Am J Clin Pathol. 2014; 142: 76-81Crossref PubMed Scopus (20) Google Scholar Due to a variety of clinical manifestation of IgG4-RD and mimicking of other diseases, there is usually a delay in diagnosis of IgG4-RD. During the workup of complex conditions, serum EPG/IFE may be requested. EPG/IFE is generally performed where symptoms suggest a monoclonal gammopathy. A monoclonal band, also known as an M protein, generally appears as homogenous, restricted, well-demarcated band in an EPG and can be further typed by IFE.4Lee A.Y.S. Cassar P.M. Johnston A.M. Adelstein S. Clinical use and interpretation of serum protein electrophoresis and adjunct assays.Br J Hosp Med. 2017; 78: C18-C20Crossref Scopus (2) Google Scholar IgG4 has a distinct pattern of migration in fast gamma region.5Finn W.G. Gulbranson R. Fisher S. et al.Detection of polyclonal increases in immunoglobulin G4 subclass by distinct patterns on capillary serum protein electrophoresis: diagnostic pitfalls and clinical observations in a study of 303 cases.Am J Clin Pathol. 2016; 146: 303-311Crossref PubMed Scopus (9) Google Scholar Normally, IgG4 serum levels are the lowest in comparison to other IgG subclasses and therefore are invisible on EPG/IFE gels which have a lower limit of detection of about 1 g/L. In the above case, a dense polyclonal IgG is noted in fast gamma region on IFE which did not appear to be homogenous or monoclonal. A monoclonal IgG4 paraprotein, on the other hand, is homogenous in appearance and has light chain restriction (Fig. 2), therefore suggesting monoclonality. In conclusion, the appearance of dense focal polyclonal IgG in fast gamma region on IFE is suggestive of elevated IgG4. When this is noted, we would suggest reflex testing of serum IgG subclasses to assist clinicians in diagnosing the possibility of IgG4-RD, and consider adjunct testing (such as serum free light chains) to investigate for possible IgG4 paraprotein. However, raised serum IgG4 is not specific for IgG4-RD and may also be found in a variety of other disorders such as malignancies, infections, autoimmune diseases, primary immunodeficiencies, idiopathic interstitial pneumonitis and vasculitides.6Ebbo M. Grados A. Bernit E. et al.Pathologies associated with serum IgG4 elevation.Int J Rheumatol. 2012; 2012: 602809Crossref PubMed Scopus (92) Google Scholar An IgG4 level of >1.35 g/L has a diagnostic sensitivity and specificity of 97% and 80%, respectively, for IgG4-RD.7Masaki Y. Kurose N. Yamamoto M. et al.Cutoff values of serum IgG4 and histopathological IgG4+ plasma cells for diagnosis of patients with IgG4-related disease.Int J Rheumatol. 2012; 2012: 580814Crossref PubMed Scopus (120) Google Scholar As a result of our observations, we now have a standardised comment that accompanies the IFE result, prompting clinicians to consider additional testing. This case report highlights the importance of the laboratory in identifying polyclonal IgG with restricted polyclonal fast gamma mobility to avoid a misdiagnosis of a monoclonal gammopathy. No funding was received for this work. The authors state that there are no conflicts of interest to disclose.