Immunophenotyping

IMMUNOPHENOTYPING

What is immunophenotyping?

Immunophenotyping is a diagnostic test that helps determine the types of proteins or markers present on/in cells. In the clinical setting it is generally utilized to further characterize certain lymphomas and leukemias.  For lymphoid tumors (such as lymphoma and lymphoid leukemias), immunophenotyping is often employed to determine if the patient’s cancer is of the B- or T-cell lineage.  This is important not only for prognostic information, but also to guide treatment recommendations in certain situations. 

Immunophenotyping can be performed in four main ways: 1) Flow cytometry of a lymph node aspirate or other organ, 2) PCR for antigen receptor rearrangement (PARR) of a lymph node aspirate or other organ, 3) immunohistochemistry (IHC) of a biopsy sample, or 4) immunocytochemistry (ICC) of an aspirate.   With flow cytometry, a fresh sample of fluid, tissue, or blood is collected, stained with specific antibodies (that recognize cellular proteins) and run through a machine that helps determine cell size, granularity, and level of protein expression.  With IHC and ICC, cells already attached to a glass slide are stained with antibodies that recognize certain cellular proteins, and positive cells turn a different color.  While IHC requires a biopsy sample (ie a piece of tissue), ICC can be performed on unstained fine needle aspirates.  PARR is a molecular procedure that evaluates cellular DNA, and determines if DNA from each cell is the same length (indicating cancer).  Reactions testing for DNA consistent with B-cell lymphoid cancer are run alongside reactions testing for T-cell lymphoid cancer for each sample.   

Which immunophenotyping procedure is preferred?

If the patient has already been diagnosed via biopsy, then IHC is the test of choice.  This test is performed on actual pieces of tissue, and can be requested by calling the pathologist and asking for special stains (eg CD3 and CD79 staining to determine if a patient has B- or T-cell lymphoma).  If the patient was diagnosed via cytology, then flow cytometry is recommended, as this will reveal the correct phenotype roughly 90% of the time.  A fresh sample is required for this test (such as blood, aspirates of lymph node/organ suspended in special media, bone marrow, or body fluids), and if this is not possible, then PARR can be performed on previously acquired lymph node or organ aspirate slides (in addition to blood, fresh or formalin fixed biopsies, bone marrow, or body fluids).  It is important to note that roughly 25% of lymphoma cases will be negative with PARR, so immunophenotype will only be revealed in 75% of cases with this test.  If PARR is negative, immunocytochemistry can be attempted using fine needle aspirates on slides that have not been previously stained.