T CELL CLONALITY (TCR GAMMA)
Label Name: TCR GAMMA
Lab Discipline: Molecular Diagnostics
Institution:  Duke University Health System 
EAP ID:  LAB6848 
Last Review:  3/17/2017 10:06:16 AM
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
TCR GAMMA
Polyclonal
Methodology
  The TCRG Gene Clonality Assay utilizes the BIOMED-2 multiplex PCR master mixes targeting the variable (V) and joining (J) regions of the rearranged TCRG gene locus to asses T-cell clonality. Genomic DNA is first extracted from the clinical sample. This DNA is then used in two independent PCR amplification reactions. One reaction amplifies DNA sequences between the V gamma segments 1-8, 10 and all J gamma segments, while the other amplifies DNA sequences between the V gamma segments 9,11 and all J gamma segments. Following PCR amplification, fluorescently labeled PCR products are resolved using capillary electrophoresis on the ABI 3130xl Genetic Analyzer. GeneMapper software (ABI) is then used to visualize and analyze PCR products, allowing detection of clonal cells representing >10% of the cell population.

This test was developed and its performance characteristics determined by this 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 TCR gamma gene locus (TCRG), mapped to 7q14, spans 160 kb and encodes the TCR gamma gene. This locus contains an extended repertoire of germline variable DNA sequences, with 14 V segments (comprised of 6 functional V segments, 3 open reading frames and 5 pseudogenes), and 5 functional joining (J) segments. The TCRG locus does not contain any D segments. Rearrangement of the TCR gamma gene locus occurs between V segments and J segments to produce a V-J TCR gamma gene. Thus, every T lymphocyte contains either one or two copies of a recombinant V-J TCR-gamma gene locus which is potentially unique in both sequence and length. During the early stages of T-cell development TCR gamma gene rearrangement follows TCR-delta gene rearrangement, with the protein products of these two genes expressed as a delta/gamma heterodimer. TCR-gamma/delta expressing cells represent a minority of T-cells, since the protein products of the delta/gamma rearrangements are only functional in a small number of cells. Following a non-functional rearrangement, the TCR-beta and TCR-alpha genes rearrange to express as an alpha/beta heterodimer in TCR-alpha/beta positive cells. Although these cells lose the expression of the TCR gamma gene product they retain the rearranged TCR gamma gene loci. Thus almost all T-cells have rearranged TCR gamma gene loci. In addition, the simple TCR gamma locus organization allows for comprehensive PCR-based amplification of nearly all gamma V-J combinations.

Like all neoplastic proliferations, T-cell lymphomas and leukemias are clonally derived. In other words, all of the cells of a T-cell neoplasm are derived from a single progenitor cell. Thus all of the neoplastic cells in a T-cell lymphoma or leukemia will have the same unique rearrangement of the TCRG locus. This property of T-cell neoplasms can be a very useful adjunct to histologic assessment, flow cytometry and immunohistochemistry in diagnosing a T-cell lymphoma or leukemia. In this assay, the distribution of sizes of the rearranged TCRG locus is determined through PCR based methods. A clonal T-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 TCRG repertoire of normal T-cells. The majority (>95%) of T-cell malignancies (including T-ALL, T-LGL, T-PLL, and MF/CTCL will test positive for TCRG clonality when using the primers sets in this assay (BIOMED 2 primers)(1,2)

References:

1. Bruggemann et. al. Powerful strategy for PCR-based clonality assessment in T-cell malignancies: - Report of the BIOMED-2 Concerted Action BHM4 CT98-3936. Leukemia (2007) 21, 215-221.

2. Van Krieken et. al. Improved reliability of lymphoma diagnostics via PCR-based clonality testing: - Report of the BIOMED-2 Concerted Action BHM4 CT98-3936. Leukemia (2007) 21, 201-206.

   
Indications
    T-cell clonality testing can be used:

As an adjunct to histologic evaluation, immunohistochemistry and flow cytometry in the diagnosis of T-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.

T-cell clonality testing may be used for:

The evaluation of residual disease after therapy. However, the sensitivity of this assay (10%) limits its utility in this regard. Please contact the Clinical Molecular Diagnostics Laboratory for other approaches to MRD testing.
   
Limitations
    This assay detects only T cell receptor gamma (TCRG) chain gene rearrangements.

A small number of TCRG rearrangements are not detectable with this assay (less than 5%).

Clonal populations that comprise less than 10% of all T cells in a sample may not be detected by this assay (analytical sensitivity of 10%).

Occasionally non-malignant processes show clonal gene rearrangements. For specimen containing a small number of T cells, psuedoclonality can occur from the preferential amplification of select DNA sequences among a limited number of amplifiable targets.

Natural killer (NK) cell lymphomas and leukemias retain the germ line configuration of the TCR gamma loci, and will test negative using this assay.

Due to the complexity of TCRG testing, results should always be interpreted in the context of clinical and pathological findings. Please consult the Clinical Molecular Diagnostics Laboratory for additional information or for help interpreting your results.
   
Test Synonyms
  Synonym(s): Leukemia
Synonym(s): Lymphoma
Synonym(s): T Cell Clonality
Synonym(s): T-Cell
Synonym(s): TCR
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