Selective immunophenotyping for diagnosis of B-cell neoplasms: immunohistochemistry and flow cytometry strategies and results

Abstract

Determining the immunophenotype of hematologic malignancies is now an indispensable part of diagnostic classification, and can help to guide therapy, or to predict clinical outcome. Diagnostic workup should be guided by morphologic findings and evaluate clinically important markers, but ideally should avoid the use of overly broad panels of immunostains that can reveal incidental findings of uncertain significance and give rise to increased costs. Here, we outline our approach to diagnosis of B-cell neoplasms, combining histologic and clinical data with tailored panels of immunophenotyping reagents, in the context of the 2008 World Health Organization classification. We present data from cases seen at our institution from 2004 through 2008 using this approach, to provide a practical reference for findings seen in daily diagnostic practice.

Figure 1

In situ pattern of follicular lymphoma. The neoplastic follicle center at the bottom labels intensely for BCL-2 whereas the germinal center of the normal secondary follicle at the top shows the typical lack of labeling. Note that the normal mantle B-cells and interfollicular T-cells show the expected pattern of staining for BCL-2. BCL-2.

Figure 2

Normal primary follicle of lymph node. Note the strong BCL-2 labeling of this primary follicle. The initial diagnosis for this lymph node was follicular lymphoma because the primary follicles were erroneously interpreted as neoplastic based on BCL-2 staining. However, these follicles lacked expression of germinal center markers, supporting their benign primary follicle nature. BCL-2.

Figure 3

Inter-follicular lymphoma cells in follicular lymphoma. In this example the HGAL labeling of inter-follicular small and large cells provides support for follicle center derivation and a diagnosis of follicular lymphoma. HGAL.

Figure 4a

Normal secondary follicle of lymph node. Note the IgM labeling in the form of immune complexes on the follicular dendritic cell processes in the pole of the germinal center adjacent to the mantle B-cells that show a circumferential (membrane-like) pattern of labeling. IgM.

Figure 4b

Neoplastic follicle. Note the intense cytoplasmic labeling of the lymphoma cells within the neoplastic follicle center in this example of follicular lymphoma that lacked expression of BCL-2 as well as CD10. IgM.

Figure 5

Lymph node involved by chronic lymphocytic leukemia. This example shows nuclear labeling for ZAP70. The residual secondary follicle at the left shows intense labeling of normal T-cells as expected. Such T-cell labeling serves as a convenient internal control. In addition, there is weak non-specific cytoplasmic labeling of some germinal center B-cells in this particular example. ZAP70.

Figure 6

In situ pattern of mantle cell lymphoma. Note the strong nuclear labeling for Cyclin D1 in the small lymphocytes in the mantle zone of this secondary follicle. Cyclin D1.

Figure 7

Marginal zone lymphoma of the lung. The lymphoma cells in this example show weak labeling for CD43. The scattered normal T-cells in this lymphoma show more intense labeling. CD43.

Figure 8

Hairy cell leukemia in the bone marrow. The hairy cells label for Annexin 1 in a membranous pattern while the residual myeloid cells show a cytoplasmic pattern of labeling. Annexin 1.

Figure 9

Multiple myeloma. The neoplastic plasma cells in this example show strong nuclear labeling for Cyclin D1. These cells also labeled for CD20 (not shown). Cyclin D1.

Figure 10

Large cell lymphoma in the setting of HIV-associated multi-centric Castleman’s disease. The large lymphoma cells show strong nuclear labeling for HHV8. HHV8.