Q & A, 9/13

Editor: Frederick L. Kiechle, MD, PhD

Submit your pathology-related question for reply by appropriate medical consultants. CAP TODAY will make every effort to answer all relevant questions. However, those questions that are not of general interest may not receive a reply. For your question to be considered, you must include your name and address; this information will be omitted if your question is published in CAP TODAY.

Submit a Question [hr] [pulledquote]Q. Our clinicians are asking about testing for IgG4-related disease. What role does IgG4 immunohistochemical staining play?
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A. IgG4-related disease is a recently recognized fibroinflammatory condition that may affect a wide variety of organ systems, producing mass lesions and generally responding to immunosuppressive therapy. The pancreas, salivary/lacrimal glands, and kidney are frequently affected, but almost any tissue may be involved, including aorta, pleura, retroperitoneum, and lymph nodes.(1-4) Several excellent reviews cover the pathophysiology, diagnostic criteria, and organ-specific features of IgG4-related disease.(1-5) Diagnosis is established based on correlation of tissue histopathologic features (dense lymphoplasmacytic infiltrate, storiform fibrosis, obliterative phlebitis ± eosinophilic infiltrate; presence of large numbers of IgG4-positive plasma cells), serologic studies (elevated serum IgG4 and/ or IgG, absent in 30 to 40 percent of cases), imaging, and clinical findings.(1-3) Indeed, the diagnosis of IgG4-related disease cannot be established with certainty without a tissue stain for IgG4.(1) However, caution is needed because the presence of IgG4-positive plasma cells in tissue is not entirely specific for IgG4-related disease. Other conditions rich in IgG4-positive plasma cells described to date include sclerosing cholangitis, ANCA-associated vasculitis, inflammatory bowel disease, sinusitis, Rosai-Dorfman disease, and atrophic gastritis, along with some lymphomas and carcinomas.(1)

Immunohistochemical staining for IgG4 is readily performed on formalin-fixed paraffin embedded sections after antigen retrieval, using the monoclonal antibody HP6025, or one of several polyclonal antibodies. Quantitative assessment of the IgG4 immunohistochemical stain with a threshold of 30 to 50 IgG4-positive plasma cells per high power field has been recommended.1 However, cut points may vary by tissue type, specimen type (core biopsy versus other), and degree of fibrosis/inflammation, with fewer plasma cells present in the predominantly fibrotic phase.1 Because of the patchy nature of involvement, needle biopsies may have false-negative results; when possible, excisional biopsies of affected tissues should be evaluated. Assessment of the IgG4:IgG plasma cell ratio has also been recommended as perhaps a more powerful method, with a proposed threshold of 40 percent in most situations.(1)

In summary, tissue immunostaining for IgG4 plays an important role in the diagnosis of IgG4-related disease, along with judicious interpretation and appropriate histologic, laboratory, and clinical correlation.

1. Deshpande V, Zen Y, Chan JK, et al. Consensus statement on the pathology of IgG4-related disease. Mod Pathol 2012;25(9):1181–1192.
2. Stone JH, Zen Y, Deshpande V. IgG4-related disease. N Engl J Med 2012;366 (6):539–551.
3. Smyrk TC. Pathological features of IgG4-related sclerosing disease. Curr Opin Rheumatol 2011;23(1):74–79.
4. Cheuk W, Yuen HK, Chu SY, et al. Lymphadenopathy of IgG4-related sclerosing disease. Am J Surg Pathol 2008;32(5):671–681.
5. Deshpande V. The pathology of IgG4-related disease: critical issues and challenges. Semin Diagn Pathol 2012;29(4):191-196.

Megan L. Troxell, MD, PhD
Associate Professor Department of Pathology
Oregon Health & Science University Portland

Member, CAP Immunohistochemistry Committee

[pulledquote]Q. We have a severely diabetic patient (glucose >800) who is a difficult draw. He has a central venous line that has insulin running in it. We have on occasion in similar circumstances asked nursing staff to shut the line off for two minutes and then obtain a 5 cc discard draw, followed by blood collected for our tests. Is the two-minute shutoff and 5 cc discard draw sufficient to ensure proper specimen integrity?[/pulledquote]

A. Based on the literature, yes, a two-minute shutoff and a 5 cc discard draw is sufficient for specimen integrity.(1) The Clinical and Laboratory Standards Institute’s “Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture” states: “Because it is normal practice to flush lines with a solution to reduce the risk of thrombosis, lines must be cleared of this fluid before blood specimens can be drawn for diagnostic testing. An adequate amount of blood must be withdrawn from the line and discarded before drawing a specimen to ensure that the actual specimen is not diluted or contaminated with the flush solution. Discard volume is dependent on the dead-space volume of the particular line. Discarding two times the dead-space volume is recommended for noncoagulation testing, and 5 mL or six times the dead-space volume for coagulation tests.”(2)

I was unable to find any references that specifically mention insulin as a deterrent to using a central line as a source for blood withdrawal. The potential risk is the direct analytical effect of insulin contamination on laboratory tests. At insulin concentrations of 3 mg/L and 1 U/L, insulin failed to induce any analytic interference when added to a variety of common chemistry tests.3 The greater clinical concern is centered around the use of heparin and the fact that the discard amount is larger for coagulation testing than for other laboratory tests.