B CELL CLONALITY (IGH)
Label Name: BCELL CL IGH
Lab Discipline: Molecular Diagnostics
Institution:  Duke University Health System 
EAP ID:  LAB6077 
Last Review:  3/16/2017 2:46:08 PM
Container & Volume
  Age Group   Container   Volume  
  0  - 18 Years LAVENDER TOP TUBE 3  ML
Label Reminders
  Be sure to include patient's name, history #, date and time of collection, and collector's initials.
Collection Notes
  All:
  • Bone Marrow: One lavender-top EDTA tube (minimum of 1 ml) is required for testing. Forward unprocessed bone marrow promptly at ambient temperatures. THE SPECIMEN CANNOT BE FROZEN. GREEN-TOP (HEPARIN) TUBES ARE NOT ACCEPTABLE FOR TESTING.

    Peripheral Blood: One lavender-top EDTA tube (minimum of 3 mls) is required for testing. Forward unprocessed peripheral blood promptly to the laboratory at ambient temperatures. THE SPECIMEN CANNOT BE FROZEN. GREEN-TOP (HEPARIN) TUBES ARE NOT ACCEPTABLE FOR TESTING.

    Fresh tissue: 200 mg tissue. Tissue should be frozen within 1 hour of collection and sent to the laboratory on dry ice. Please contact the Clinical Molecular Diagnostics Laboratory for help shipping samples.

    Formalin Fixed Paraffin Embedded Tissue: The laboratory can receive either a paraffin embedded tissue block or four freshly cut (within one week)5uM thick unstained slides containing 3 to 20 square mm of tissue. Unstained slides should be accompanied by an H&E stained slide for histologic evaluation.

    Fine Needle Aspiration Samples: For internal specimens, a fine needle aspirate sample should be sent via courier in a 15 ml conical tube containing RPMI. Forward specimen promptly at ambient temperature. DO NOT FREEZE.

    Cerebrospinal fluid (CSF): Forward unprocessed CSF promptly at ambient temperatures. THE SPECIMEN CANNOT BE FROZEN.
 
Transport
  Deliver peripheral blood and bone marrow specimens and FNA specimens to laboratory at ambient temperature. If there is a delay of more than 24 hours in delivery, refrigerate the sample. DO NOT FREEZE. Tissue should be sent frozen on dry ice.
Turn Around Time -  Routine: 10 days   Stat: N/A
Reference Values
BCELL CL IGH
NEGATIVE
Methodology
  This assay uses PCR targeting the variable (V) and joining (J) regions of the rearranged IGH gene locus to assess B-cell clonality. Genomic DNA is first extracted from the clinical sample. This DNA is then used in two independent PCR amplification reactions. One reaction targets framework 1 sequences within the V region, while the other targets framework 3. Following PCR amplification, fluorescently labeled PCR products are resolved by capillary electrophoresis on the ABI 310 Genetic Analyzer. GeneScan software (ABI) is then used to visualize and analyze PCR products, allowing detection of clonal populations comprising >5% of all B cells in the sample.

This test was developed and its performance characteristics determined by the DUHS Clinical Molecular Diagnostics Laboratory. It has not been cleared or approved by the U.S. Food and Drug Administration. This test is used for clinical purposes. It should not be regarded as investigational or for research. This laboratory is certified under the Clinical Laboratory Improvement Amendments of 1988 ("CLIA") as qualified to perform high complexity clinical testing.
   
Special Information
  Adult:
  • Additional Patient Information Required: Send all pertinent clinical information with specimen.
 
   
Clinical Significance and Interpretive Data
    The human immunoglobulin heavy chain locus (IGH), mapped to 14q32.3, spans 1250 kb and encodes the immunoglobulin heavy chain. This locus contains an extended repertoire of germline variable DNA sequences, with approximately 52 functional V segments, 27 functional diversity (D) segments, and 6 functional joining (J) segments. The heavy chain V segments contain three framework regions (FR1, FR2 and FR3), and two complementarity-determining regions (CDR1 and CDR2). Immature B-cells undergo somatic recombination that joins individual D and H segments, followed by V segment rearrangement, creating a functional V-D-J IgH gene. This can happen in either or both of the IgH loci. Thus, every B lymphocyte contains either one or two copies of a recombinant V-D-J heavy chain which is potentially unique in both sequence and length.

Like all neoplastic proliferations, B-cell lymphomas and leukemias are clonally derived. In other words, all of the cells of a B-cell neoplasm are derived from a single progenitor cell. Thus all of the neoplastic cells in a B-cell lymphoma or leukemia will have the same unique rearrangement of the IGH locus. This property of B-cell neoplasms can be a very useful adjunct to histologic assessment, flow cytometry and immunohistochemistry in diagnosing a B-cell lymphoma or leukemia. In this assay, the distribution of sizes of the rearranged IGH locus is determined through PCR based methods. A clonal B-cell population manifests as an overrepresentation of a specific length PCR amplification product. This provides a detectable molecular marker which is distinguishable from the diverse IGH repertoire of normal B-cells.
   
Indications
    B-cell clonality testing can be used:

As an adjunct to histologic evaluation, immunohistochemistry and flow cytometry in the diagnosis of B-cell lymphomas and leukemias

To determine if two samples contain the same clonal or neoplastic process to help distinguish recurrent or residual disease from reactive inflammation or a new lymphoid neoplasm.

B-cell clonality testing may be used for:

The evaluation of residual disease after therapy. However, the sensitivity of this assay (5%) limits its utility in this regard. Please contact the Clinical Molecular Diagnostics Laboratory for other approaches to MRD testing.

   
Limitations
    This assay detects the B-cell heavy chain gene rearrangements only.

Clonal populations that comprise less than 5% of all B-cells in a sample may not be detected (analytical sensitivity of 5%).

Greater than 80% of IGH V-D-J rearrangements will be detected by this assay. However, due to the potential DNA sequence diversity of these rearranged IGH loci, not all will be amplifiable by our PCR primer sets. Thus, a negative result does not preclude the possibility of a clonal B-cell proliferation.

Occasionally non-malignant processes show clonal gene rearrangements. For specimen containing a small number of B cells, psuedoclonality can occur from the preferential amplification of select DNA sequences among a limited number of amplifiable targets.
   
Test Synonyms
  Synonym(s): B-cell
Synonym(s): Leukemia
Synonym(s): Lymphoma
Molecular Diagnostics Laboratory
(MDX)

Medical Director:
 Michael Datto, M.D., Ph.D.
 Phone: 919-684-6965
 Email: michael.datto@duke.edu
Lab Director:
 Catherine Rehder Ph.D, FACMG
 Phone: 919-613-8434
 Email: catherine.rehder@duke.edu
Lab Director:
 Siby Sebastian Ph.D., DABMG
 Phone: 919-613-8432
 Email: siby.s@duke.edu

Address: 
 Wadsworth Bldg, Cytogenetics, Rm 0220
 2351 Erwin Rd
 Durham,  NC  27705
 Phone: 919-684-2698
 FAX: 919-668-5424

Performing Times:
  Monday through Friday