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Immunophenotyping with Canine Lymphoma | VETgirl Veterinary CE Blog

Immunophenotyping:
In veterinary medicine, currently canine lymphoma is broken down into two main immunophenotypes, B and T cell. Traditionally, 2/3 of dogs with lymphoma are classified as B cell and 1/3 are T cell. A minor percentage (<2%) are deemed null cell. Immunophenotyping of lymphoma patients can be achieved through a variety of tests including immunohistochemistry, immunocytochemistry, polymerase chain reaction (PCR), and flow cytometry.1

  • Immunohistochemistry (IHC):  IHC is still considered the gold standard for determining immunophenotype, utilizing a panel of markers that bind to surface proteins either on B cells (CD79a, CD21, Pax5) or T cells (CD5, CD3, CD4, CD8). This requires tissue obtained either via a punch biopsy, tru-cut biopsy, or nodal extirpation.
  • Immunocytochemistry (ICC): ICC utilizes the same antibodies as IHC but on cytology samples, thus offering a more cost effective manner for which to determine immunophenotype. The distinction is that nodal architecture is not evaluated, thus the grade and specific subtype (e.g. indolent T zone, diffuse large B cell, etc) of lymphoma cannot be determined.
  • Polymerase Chain Reaction (PCR): This is a repetitive enzymatic reaction that generates ~109 copies of a particular DNA sequence from 1 original copy, thus a small sample can yield diagnostic results. It utilizes heat-stable polymerases and sequence-specific primers.  This test is commonly used in the detection of infectious disease in human and veterinary medicine. PCR for antigen receptor rearrangement (PARR):  Clonality is the hallmark of malignancy, and PARR amplifies the variable regions of immunoglobulin genes and T cell receptor genes to detect the presence of a clonal population. PARR not only confirms clonality, and thus cancer, but will also determine the immunophenotype of lymphoma and lymphoid leukemia. Specific samples that can be analyzed include: peripheral blood, lymph node, bone marrow, mediastinal mass, body cavity fluid, and cerebral spinal fluid.1
  • Flow Cytometry (FCM):  FCM is routinely used in physician-based medicine early in the diagnostic work-up of lymphoid malignancies and involves the use of fluorescently-labeled monoclonal antibodies. This allows for the evaluation of a large population of cells in order to determine cell sizes, identify aberrant surface marker expression, and thus determine the immunophenotype of both normal and atypical cells in the sample., FCM requires fresh samples (live cells) and is commercially available through several major diagnostic laboratories.1

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