IGH Clonality Assays

• Description of Test:

  For detection of the vast majority of clonal IGH V_H-J_H rearrangements, including the associated V_H-J_H region DNA sequences, a multiplex master mix targeting the conserved framework region 1 (FR1), framework region 2 (FR2), or framework region 3 (FR3), as well as the joining region, is used for PCR amplification.  Next-generation sequencing of the PCR products is used to identify DNA sequences specific to clonal gene rearrangements.  Bioinformatics tools facilitate the characterization of sequences present at greater than 2.5% of the population.  These sequences can be used to track specific clonal populations.

  Test Names
  IGH FR1 Clonality Assay
  IGH FR2 Clonality Assay
  IGH FR3 Clonality Assay

• Overview:

  Lymphoid cells are different from the other somatic cells in the body as during development, the antigen receptor genes of these cells undergo somatic gene rearrangement.¹

  The human immunoglobulin heavy chain (IGH) gene locus on chromosome 14 (14q32.3) includes 46-52 functional and 30 non-functional variable (V_H) gene segments, 27 functional diversity (D_H) gene segments, and 6 functional joining (J_H) gene segments spread over 1250 kilobases.  During B-cell development, genes encoding the IGH molecules are assembled from multiple polymorphic gene segments that undergo rearrangements and selection.  These gene rearrangements of the variable, diversity and joining segments generate V_H-D_H-J_H combinations of unique length and sequence for each cell.^2,3

  Since leukemia and lymphomas originate from the malignant transformation of individual lymphoid cells, all leukemias and lymphomas generally share one or more cell-specific or “clonal” antigen receptor gene rearrangements.  Clonality does not always imply malignancy; all results must be interpreted in the context of all of the other available diagnostic criteria.  Tests that detect IGH clonal rearrangements are useful in the characterization, monitoring, and treatment of B- and T-cell malignancies.

   

  CLINICAL UTILITY

  Immunoglobulin heavy chain (IGH) clonality studies are often used in practice in conjunction with morphologic and immunophenotypic evaluation to distinguish B-cell neoplasm from reactive B-cell proliferation or reactive plasmacytosis.  They are particularly useful under the following conditions:

  •   A suspected/equivocal clonal B-cell population is detected by flow cytometry, and clonality needs to be confirmed;

  •   Atypical lymphoid infiltrate is present on the tissue sections (lymph nodes or extra nodal tissue) sections based on morphology review, and immunophenotypic studies by flow cytometry are not available and/or no obvious immunophenotypic aberrancy can be demonstrated by immunohistochemical stains;

  •   Dense plasmocytic infiltrate on the tissue sections (lymph nodes, bone marrow clot or other extra nodal tissue) based on morphology review, and

  −  immunophenotypic studies by flow cytometry are not available to evaluate the plasma cells and/or B-cells,

  −  no obvious clonal plasma cell population can be demonstrated by immunohistochemical stains or in situ hybridization, and

  −  no obvious immunophenotypic aberrancy can be demonstrated on the plasma cells and/or B-cells by immunohistochemical stains;

  •   Investigating the clonal relationship between the current specimen with a diagnosis of B-cell lymphoma/leukemia and a prior specimen with a diagnosis of B-cell lymphoma/leukemia.

  While IGH clonality studies are essential for the differential diagnosis of B-cell lymphoma vs. reactive B-cell proliferation sometimes, it is important to understand that the diagnosis cannot be based on molecular studies alone but requires careful correlation of clinical, morphologic, immunophenotypic and genetic features.  IGH clonality studies may be performed in conjunction with immunoglobulin kappa light chain (IGK) clonality studies.

Service Details

• Indications for Testing:
  • Identify clonality in atypical lymphoproliferative disorders
  • Support a differential diagnosis between reactive lesions and hematologic malignancies
  • Assign presumptive lineage in mature monoclonal lymphoproliferative disorders
  • Monitor and evaluate disease recurrence
• Service Description: