[1766] Automated Double Immunoenzymatic and Colorimetric In-Situ Hybridization Techniques in Evaluation of Immunoglobulin Light Chain Expression

BA Webber, C Cohen. Atlanta VAMC, Decatur, GA; Emory University, Atlanta, GA

Background: In diagnosis of lymphoid/plasma cell lesions, when flow cytometry results are unavailable, it may be necessary to assess paraffin-embedded fixed tissues to establish B cell/plasma cell clonality. Single color detection based immunohistochemistry for kappa and lambda immunoglobulin light chains is complicated by high background staining, poor sensitivity in non-plasma cell populations, and difficulty identifying particular cell populations in adjacent sections. We compared two commercially available alternative automated methods for evaluating kappa/lambda expression, a double immunoenzymatic (DIE) technique and in-situ hybridization (ISH), in tissues harboring lymphoid or plasma cell lesions processed with B5 (both decalcified and non-decalcified), formalin, and B-plus fixatives.
Design: Single sections of archived paraffin embedded tissues (Emory University, Atlanta, GA) were stained via automated DIE using monoclonal antibodies for kappa and lambda on a Bondmax (Leica) Autostainer and compared with adjacent serial sections stained via automated ISH using pre-diluted kappa and lambda probes on a Ventana Benchmark automated stainer. Cases included 17 plasma cell dyscrasia/myeloma marrows, 1 soft tissue plasmacytoma, 3 diffuse large B cell lymphomas, 2 follicular lymphomas, 6 marginal zone lymphomas, 1 Castleman disease, 1 benign marrow plasmacytosis, and 18 tonsillar lymphoid hyperplasias. All marrows were B5-fixed and decalcified using a hydrochloric acid (HCl) based rapid decalcifying solution, while the remaining cases were processed using formalin, B5, or B-plus fixative. Findings were correlated with previously documented flow cytometric immunophenotyping results.
Results: Correct assessment of clonality was achieved in 33 (66%) and in 28 of 50 (56%) cases by DIE and ISH, respectively. The remaining cases were indeterminate because of absent or poor quality staining. Kappa/lambda expression was only observed in plasma cells, with no reliable staining of lymphoid populations identified by either method. While fixation method had no adverse effect, a large proportion of decalcified specimens yielded non-diagnostic results because of poor quality or absent staining.
Conclusions: DIE and ISH overall are moderately effective methods for evaluating immunoglobulin light chain expression in plasma cells, but these methods demonstrate only limited success in tissues undergoing rapid, HCl-based decalcification.
Category: Techniques

Wednesday, March 11, 2009 1:00 PM

Poster Session VI # 246, Wednesday Afternoon