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Test Code MISC2MAYOTREC T-Cell Receptor Excision Circles (TREC) Analysis, Blood

Important Note

Collect Monday-Thursday ONLY.

Reporting Name


Useful For

Measuring T-cell output or reconstitution (thymopoiesis) following hematopoietic cell transplantation or highly active antiretroviral therapy


Evaluating thymic function in patients with cellular or combined primary immunodeficiencies, or receiving immunotherapy or cancer vaccines


Assessing T-cell recovery following thymus transplants for DiGeorge syndrome

Performing Laboratory

Mayo Clinic Laboratories in Rochester

Specimen Type

Whole Blood EDTA

Additional Testing Requirements

This assay is useful for longitudinal assessment of thymic function and naive T-cell production. For comprehensive thymic function analysis on pediatric patients or posthematopoietic cell transplantation, order this test and CD4RT / CD4 T-Cell Recent Thymic Emigrants (RTE).

Shipping Instructions

Specimens are required to be received in the laboratory weekdays and by 4 p.m. on Friday. Draw and package specimen as close to shipping time as possible.


It is recommended that specimens arrive within 24 hours of draw.


Samples arriving on the weekend and observed holidays may be canceled.

Necessary Information

Ordering physician's name and phone number are required.

Specimen Required

For serial monitoring, we recommend that specimen draws be performed at the same time of day.


Supplies: Ambient Shipping Box-Critical Specimens Only (T668)

Container/Tube: Lavender top (EDTA)

Specimen Volume:

Adults: 10 mL


-Preferred volume for >1 year = 5 mL

-Preferred volume for ≤1 year old = 3 mL

Collection Instructions:

1. Do not draw specimen through a butterfly needle.

2. Send specimen in original tube. Do not aliquot.

Specimen Minimum Volume

Adults: 10 mL/Pediatrics: 1 mL

Specimen Stability Information

Specimen Type Temperature Time Special Container
Whole Blood EDTA Ambient 48 hours PURPLE OR PINK TOP/EDTA

Special Instructions

Reference Values

The appropriate age-related reference values will be provided on the report.

Day(s) Performed


Test Classification

This test was developed, and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the US Food and Drug Administration.

CPT Code Information

81479-Unlisted molecular pathology procedure

LOINC Code Information

Test ID Test Order Name Order LOINC Value
TREC TREC, B In Process


Result ID Test Result Name Result LOINC Value
28306 CD3 T Cells 8122-4
28307 CD4 T Cells 24467-3
28308 CD8 T Cells 14135-8
87959 TREC Copies 62320-7
30292 Interpretation 69047-9
35200 Reviewed by 18771-6

Clinical Information

T cell reconstitution is a critical feature of the recovery of the adaptive immune response and has 2 main components: thymic output of new T cells and peripheral homeostatic expansion of preexisting T cells. It has been shown that though thymic function declines with age, a reasonable output is still maintained into late adult life.(1) In many clinical situations, thymic output is crucial to the maintenance and competence of the T cell effector immune response.


Thymic function can be determined by T-cell receptor excision circle (TREC) analysis. TRECs are extrachromosomal DNA byproducts of T-cell receptor (TCR) rearrangement, which are nonreplicative. TRECs are expressed only in T cells of thymic origin and each cell is thought to contain a single copy of TREC. Hence, TREC analysis provides a very specific assessment of T-cell recovery (eg, after hematopoietic cell transplantation) or numerical T-cell competence. There are several TRECs generated during the process of TCR rearrangement and the TCR delta deletion TREC (deltaREC psi-J-alpha signal joint TREC) has been shown to be the most accurate TREC for measuring thymic output.(2) This assay measures this specific TREC using quantitative, real-time PCR.


Clinical use of TRECs in HIV and Antiretroviral Therapy:

HIV infection leads to a decrease in thymic function. Adult patients treated with highly active antiretroviral therapy (HAART) show a rapid and sustained increase in thymic output.(1)


Clinical use of TRECs in Hematopoietic Cell Transplantation (HCT) and Primary Immunodeficiencies (PID):

Following HCT, there is a period of prolonged immunodeficiency that varies depending on the nature and type of stem cell graft used and the conditioning regimen, among other factors. This secondary immunodeficiency also includes defects in thymopoiesis.(3-5) It has been shown that numerical T cell recovery is usually achieved by day 100 posttransplant, though there is an inversion of the CD4:CD8 ratio that can persist for up to a year.(4) Also, recovery of T-cell function and diversity can take up to 12 months, although this can be more rapid in pediatric patients. However, recovery of T-cell function is only possible when there is numerical reconstitution of T cells. T cells, along with the other components of adaptive immunity, are key players in the successful response to vaccination post-HCT.(6)


Recently, it has been shown in patients who received HCT for severe combined immunodeficiency (SCID) that T cell recovery early after stem cell transplant is crucial to long-term T cell reconstitution.(7) Patients who demonstrated impaired reconstitution were shown to have poor early grafting, as opposed to immune failure caused by accelerated loss of thymic output or long-term graft failure. In this study, the numbers of TRECs early after HCT were most predictive for long-term reconstitution. This data suggests that frequent monitoring of T-cell immunity and TREC numbers after HCT can help identify patients who will fail to reconstitute properly, which would allow additional therapies to be instituted in a timely manner.(7) It would be reasonable to extrapolate such a conclusion to other diseases that are also treated by HCT.


TREC Copies and Thymic Output in Adults:

Since the adult thymus involutes after puberty and is progressively replaced by fat with age, thymus-dependent T cell recovery has been assumed to be severely limited in adults. However, with TREC analysis it has been shown that the change in thymic function in adults is a quantitative phenomenon rather than a qualitative one and thymic output is not totally eliminated.(1,8,9) Thus, after HCT or HAART, the remaining thymic tissue can be mobilized in adults to replenish depleted immune systems with a potentially broader repertoire of naive T cells. Douek et al have shown that there is a significant contribution by the thymus to immune reconstitution after myeloablative chemotherapy and HCT in adults.(8) In fact, this data shows that there is both a marked increase in the TREC numbers and a significant negative correlation of TREC copies with age posttransplant.


In addition to the specific clinical situations elucidated above, TREC analysis can be helpful in identifying patients with primary immunodeficiencies and assessing their numerical T-cell immune competence. It can also be used as a measure of immune competence in patients receiving immunotherapy or cancer vaccines, where maintenance of, T-cell outputis integral to the immune response against cancer.


The absolute counts of lymphocyte subsets are known to be influenced by a variety of biological factors, including hormones, the environment, and temperature. The studies on diurnal (circadian) variation in lymphocyte counts have demonstrated progressive increase in CD4 T-cell count throughout the day, while CD8 T cells and CD19+ B cells increase between 8:30 am and noon, with no change between noon and afternoon. Natural killer (NK) cell counts, on the other hand, are constant throughout the day.(10) Circadian variations in circulating T-cell counts have been shown to be negatively correlated with plasma cortisol concentration.(11-13) In fact, cortisol and catecholamine concentrations control distribution and, therefore, numbers of naive versus effector CD4 and CD8 T cells.(11) It is generally accepted that lower CD4 T-cell counts are seen in the morning compared with the evening,(14) and during summer compared to winter.(15) These data, therefore, indicate that timing and consistency in timing of blood collection are critical when serially monitoring patients for lymphocyte subsets.


T-cell receptor excision circles (TRECs) generally show an inverse correlation with age, though there can be substantial variations in TREC copies relative to T-cell count within a given age group.


Following hematopoietic cell transplantation (HCT), highly active antiretroviral therapy (HAART), thymic transplants, etc, TREC typically increases from absent or very low levels (below age-matched reference range) to baseline levels or exceeds baseline levels, showing evidence of thymic rebound, which is consistent with recovery of thymic output and T-cell reconstitution.


When a patient is being monitored for thymic recovery posttransplant treatment, this assay recommends that a pretransplant (prior to myeloablative or nonmyeloablative conditioning) or a pretreatment baseline specimen be provided so that appropriate comparisons can be made between the pre- and posttransplant treatment specimens. Since there is substantial variability between individuals in TREC copies, the best comparison is made to the patient's own baseline specimen rather than the reference range (which provides a guideline for TREC copies for age-matched healthy controls).


A consultative report will be generated for each patient.

Method Description

This assay involves both preanalytical preparation of a pure cell population followed by analytical evaluation of the DNA. A modified peripheral blood mononuclear cells (PBMCs) isolation is used to prepare a nearly pure population of CD3+ T cells (adults) or total lymphocytes (pediatrics) from whole blood. The resulting purity and cell counts are obtained from the TCD4 flow cytometric assay. The cells are then lysed with Proteinase K to a predetermined target concentration, to release and expose the DNA for PCR. The genomic DNA and TREC in the cell lysates are quantified in the real-time PCR assay, in triplicate, by using a fluorescent probe specific for the TCR delta-deletion T-cell receptor excision circle (TREC) signal joint and a distinct fluorescent probe for the housekeeping gene, albumin. There is 1 copy of TREC per CD3+ T cell, while there are 2 copies of albumin in every cell. A standard curve is used to determine the absolute quantity of TREC and albumin from the fluorescence intensities measured. The albumin counts are used to determine the cell counts in each reaction and to normalize the number of TREC copies to a standard reporting unit of copies per million CD3+ T cells. The pediatric TREC counts, though measured from total lymphocytes, can be adjusted to the same reporting units using the %CD3 purity from the flow cytometric assay.(Douek DC, Vescio RA, Betts MR, et al: Assessment of thymic output in adults after hematopoietic stem cell transplantation and prediction of T cell reconstitution. Lancet 2000;355:1875-1881; Douek DC, Hill B: Personal Communication, 2005)

Report Available

6 to 8 days

Reject Due To

Gross hemolysis Reject
Gross lipemia Reject

Method Name

Real-Time Quantitative Polymerase Chain Reaction (PCR)


TREC Assay Patient Information (T589) in Special Instructions

Secondary ID