Application of Immunophenotyping and Heteroduplex Polymerase Chain Reaction (hPARR) for Diagnosis of Canine Lymphomas

Abstract

Background: Canine malignant lymphoma is classified into B- or T-cell origin, as in the human case. Due to differences in prognosis, a suitable method needs to be developed for lineage identification.

Aims: To determine the accuracy of immunophenotypic and molecular information between three methods: immunocytochemistry (ICC), immunohistochemistry (IHC) and heteroduplex polymerase chain reaction for antigen receptor rearrangements (hPARR) in spontaneous canine lymphomas.

Materials and methods: Peripheral blood, fine needle aspiration and tissue biopsies from enlarged peripheral lymph nodes prior to treatment of 28 multicentric lymphoma patients were collected. Cytopathology and histopathology were examined and classified using the updated Kiel and WHO classifications, respectively. Anti-Pax5 and anti-CD3 antibodies as B- and T-cell markers were applied for immunophenotyping by ICC and IHC. Neoplastic lymphocytes from lymph node and white blood cell pellets from peripheral blood were evaluated by hPARR.

Results: In this study, low grade B-cell lymphoma accounted for 25% (7/28), high grade B-cell lymphoma for 64.3% (18/28) and high grade T-cell lymphoma for 10.7% (3/28). According to the WHO classification, 50% of all cases were classified as diffuse large B-cell lymphoma. In addition, ICC showed concordant results with IHC; all B-cell lymphomas showed Pax5+/CD3, and all T-cell lymphomas exhibited Pax5-/CD3+. In contrast to hPARR, 12 B-cell lymphomas featured the IgH gene; seven presented the TCRγ gene; five cases showed both IgH and TCRγ genes, and one case were indeterminate. Three T-cell lymphomas showed the TCRγ gene. The percentage agreement between hPARR and ICC/IHC was 60%.

Conclusions: Immunophenotyping should not rely on a single method. ICC or IHC with hPARR should be used concurrently for immunophenotypic diagnosis in canine lymphomas.