Automated Dual Color Kappa/Lambda mRNA In Situ Hybridization for Detection of Monoclonality Using Routine Light Microscopy in B-Cell Non-Hodgkin Lymphoma
Sarah McGinn, William Day, Anne Pedata, Christopher Morrison, Thomas Grogan, Lisa Rimsza. University of Arizona, Tucson, AZ; Ventana Medical Systems, Inc., Tucson, AZ
Background: B-cell non-Hodgkin lymphomas (B-NHL) arise from monoclonal proliferation of B cells which can be detected by sole expression of either kappa or lambda light chain mRNA and protein as part of their B cell receptor antibody. Uniform expression of either light chain by malignant B cells enables differentiation of monoclonal B cell lymphomas from polyclonal mixed kappa and lambda light chain expressing B cell populations that result during the normal immune response. Determination of light chain expression patterns is complicated by the copy number range of light chain mRNA and antibody protein expressed by B cell neoplasms derived from a variety of B cell stages (naïve and memory cells: 10-100 mRNA copies per cell; plasma cells: approximately 100,000 mRNA copies per cell).
Design: In this project, we evaluated the accuracy of an automated, dual color, colorimetric, in situ hybridization (CISH) assay for simultaneous detection of kappa and lambda light chain mRNA in FFPE tissue as compared to flow cytomety. We assessed 20 B-NHL samples representing different maturational stages including 7 diffuse large B cell lymphomas, 10 follicular lymphomas, 1 MALT lymphoma, 1 nodal marginal zone lymphoma, and 1 small lymphocytic lymphoma and 1 reactive tonsil. Staining results were evaluated by 2 pathologists (SM and LR) and compared to the flow cytometry results.
Results: The dual color CISH assay detected both kappa and lambda mRNA in a reactive tonsil in a polyclonal pattern in the germinal centers, mantle zones, scattered small and large paracortical B cells, and plasma cells. While the plasma cells demonstrated abundant mRNA, the small B cells exhibited one to several small perinuclear dots per cell. The overall agreement between the CISH and the flow cytometry results was 18/20 (90%). The 2 misinterpretations were both follicular lymphomas, which were interpreted as polyclonal by CISH however, demonstrated kappa light chain restriction by flow cytometry
Conclusions: In summary, evaluation of monoclonality in lymphoma samples is feasible using the two-color CISH detection system. The assay's in situ analyses and low level of detection may allow determination of light chain clonality in a variety of B cell lymphomas using routine light microscopy on a single glass slide.
Tuesday, March 5, 2013 1:00 PM
Poster Session IV # 266, Tuesday Afternoon