Test Catalog

Test Id : TREGS

T-Cell Subsets, Regulatory (Tregs), Blood

Useful For
Suggests clinical disorders or settings where the test may be helpful

Evaluating patients with clinical features of IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked inheritance) and other primary immunodeficiencies, autoimmune diseases, allergy and asthma, and graft-vs-host disease post-hematopoietic stem cell transplantation

Method Name
A short description of the method used to perform the test

Flow Cytometry

NY State Available
Indicates the status of NY State approval and if the test is orderable for NY State clients.

No

Reporting Name
Lists a shorter or abbreviated version of the Published Name for a test

T Cell Subsets, Regulatory (Tregs)

Aliases
Lists additional common names for a test, as an aid in searching

Allograft rejection

Autoimmunity

FOXP3 mutation

FOXP3+ T-cells

GVHD

IPEX

Natural naive Tregs

Natural Tregs

Regulatory T-cells

STAT5b mutation

Tr1/Th3 T-cells

Tregs

Specimen Type
Describes the specimen type validated for testing

Whole Blood EDTA

Shipping Instructions

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

 

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

 

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

Necessary Information

Ordering physician's name and phone number are required.

Specimen Required
Defines the optimal specimen required to perform the test and the preferred volume to complete testing

Container/Tube: Lavender top (EDTA)

Specimen Volume: 3 mL

Collection Instructions: Send specimen in original tube. Do not aliquot.

Additional Information: For serial monitoring, it is recommended that specimens are collected at the same time of day.

Specimen Minimum Volume
Defines the amount of sample necessary to provide a clinically relevant result as determined by the Testing Laboratory

1 mL

Reject Due To
Identifies specimen types and conditions that may cause the specimen to be rejected

Gross hemolysis Reject
Gross lipemia Reject

Specimen Stability Information
Provides a description of the temperatures required to transport a specimen to the performing laboratory, alternate acceptable temperatures are also included

Specimen Type Temperature Time Special Container
Whole Blood EDTA Ambient (preferred) 72 hours PURPLE OR PINK TOP/EDTA

Useful For
Suggests clinical disorders or settings where the test may be helpful

Evaluating patients with clinical features of IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked inheritance) and other primary immunodeficiencies, autoimmune diseases, allergy and asthma, and graft-vs-host disease post-hematopoietic stem cell transplantation

Clinical Information
Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test

Regulatory T cells (Tregs) are a population of CD4+ T cells with a unique role in the immune response. Tregs are crucial in suppressing aberrant pathological immune responses in autoimmune diseases, transplantation, and graft-vs-host disease after allogeneic hematopoietic stem cell transplantation.(1) Tregs are activated through the specific T-cell receptor, but their effector function is nonspecific, and they regulate the local inflammatory response through cell-to-cell contact and cytokine secretion.(2) Tregs secrete interleukin (IL)-9, IL-10, and transforming growth factor-beta 1 (TGF-beta 1), which aid in the mediation of immunosuppressive activity.

 

Chief characteristics of the Treg population are surface expression of the CD25 protein (IL-2Ra) and the intracellular presence of the transcription factor FOXP3. The IL-7 receptor (CD127) is downregulated on FOXP3+CD4+CD25+ T cells and provides an excellent alternative cell-surface marker to FOXP3 for detecting natural Tregs (CD4+CD25+CD127lo).(2)

 

Natural Tregs account for 5% to 10% of the total CD4 T-cell population and are derived from thymic precursors.(3) Since CD25 is also expressed on activated T cells, the concomitant use of CD127 permits the differentiation of Tregs from activated T cells.(4) Natural Tregs express the memory marker CD45RO and have limited ability to proliferate. However, within the CD4+CD25+Treg population, there is a subset of Tregs that express the CD45 isoform generally associated with naive T cells (CD45RA), and this subset has been called natural naive (Nn) Tregs. Nn Tregs are most prominent in young adults and decrease with age along with the rest of the naive CD4 T-cell population.(5) Like other naive T cells, Nn Tregs have high proliferative capacity, as well as the suppressor activity of other Treg subsets. Evidence suggests that Nn Tregs also have a thymic ancestry and are the precursors of the natural Tregs (that are of the memory, antigen-experienced phenotype) and appear to be composed of T cells with self-reactive T-cell receptors.(5)

 

Other subsets of Tregs include the T helper 3 (Th3) cells, which secrete high levels of TGF-beta 1 and can be induced by oral administration of antigen, and regulatory T class 1 (Tr1) cells, which secrete interferon-gamma and IL-10.(5) These Treg subsets are most likely induced in the periphery and are responsible for peripheral tolerance to self-antigens. The suppressive activity of Th3 and Tr1 cells are related to the cytokines they produce, TGF-beta 1 and IL-10, respectively.

 

The absence of Tregs as a result of variants in the FOXP3 gene causes a primary immunodeficiency called IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked inheritance).(6) Patients with IPEX have a complex manifestation of symptoms including severe watery diarrhea due to significant villous atrophy and lymphocytic infiltration of bowel mucosa, early-onset autoimmune endocrinopathies involving the pancreas or thyroid, and a dermatitic (eczematous) rash. In addition, there are other autoimmune manifestations including autoimmune cytopenias and autoimmune hepatitis. Kidney disease is quite common in these patients. Finally, these patients also have a significant predisposition to infections including sepsis, pneumonia, meningitis, and osteomyelitis.(6) Decreased FOXP3+CD4+CD25+Tregs have been reported in 1 patient with a STAT5b alteration.(7)

 

There is an expansion of Nn Tregs in patients with monoclonal gammopathy of undetermined significance and multiple myeloma, likely as a response to the process of malignant transformation.(8) Expansion of Tregs has also been reported in other neoplasias including B-cell chronic lymphocytic leukemia, Hodgkin disease, and solid tumors.

 

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 cell counts, on the other hand, are constant throughout the day.(9) Circadian variations in circulating T-cell counts have been shown to negatively correlate with plasma cortisol concentration.(10-12) In fact, cortisol and catecholamine concentrations control distribution and, therefore, numbers of naive versus effector CD4 and CD8 T cells.(10) It is generally accepted that lower CD4 T-cell counts are seen in the morning compared with the evening,(13) and during summer compared to winter.(14) These data, therefore, indicate that timing and consistency in timing of blood collection is critical when serially monitoring patients for lymphocyte subsets.

Reference Values
Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.

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

Interpretation
Provides information to assist in interpretation of the test results

The lack of regulatory T cells (Tregs) is associated with variants in the FOXP3 gene. Low Tregs are also seen in the context of STAT5b alterations. Reduced Nn Tregs and natural Tregs are likely to predispose to autoimmunity, while reductions in Th3/Tr1 cells may impair oral and peripheral tolerance, also facilitating the development of autoimmunity.

 

The presence of expanded naive Tregs may indicate a process of malignant transformation, if other clinical features of malignant disease are present.

 

Increased Tregs in donor stem cell allografts have been associated with a reduced incidence of graft-versus-host disease (ie, mediating a protective effect) after allogeneic stem cell transplantation.

Cautions
Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances

This panel provides only quantitative information regarding the various regulatory T cell (Treg) subsets; it does not provide information on the functional aspect of these populations.

 

Results should be correlated with clinical presentation.

 

Molecular testing is required to confirm a diagnosis of IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked inheritance). Call 800-533-1710 for assistance in ordering molecular testing.

 

Treg cells may be reduced in a variety of clinical contexts such as in autoimmune diseases or allograft rejection.

 

Timing and consistency in timing of blood collection is critical when serially monitoring patients for lymphocyte subsets. See data under Clinical Information.

Clinical Reference
Recommendations for in-depth reading of a clinical nature

1. Sakaguchi S, Sakaguchi N, Shimizu J, et al: Immunologic tolerance maintained by CD25+CD4+ regulatory T cells: their common role in controlling autoimmunity, tumor immunity, and transplantation tolerance. Immunol Rev. 2001 Aug;182:18-32

2. Liu W, Putnam AL, Xu-Yu Z, et al: CD127 expression inversely correlates with FOXP3 and suppressive function of human CD4+ Treg cells. J Exp Med. 2006 Jul 10;203(7):1701-1711

3. Seddiki N, Santner-Nanan B, Tangye SG, et al: Persistence of naive CD45RA+ regulatory T-cells in adult life. Blood. 2006 Apr 1;107(7):2830-2838

4. Seddiki N, Santner-Nanan B, Martinson J, et al: Expression of interleukin (IL)-2 and IL-7 receptors discriminates between human regulatory and activated T-cells. J Exp Med. 2006 Jul 10;203(7):1693-1700

5. Valmori D, Merlo A, Souleimanian NE, Hesdorffer CS, Ayyoub M: A peripheral circulating compartment of natural naive CD4 Tregs. J Clin Invest. 2005 Jul;115(7):1953-1962

6. Torgerson TR, Ochs HD: Immune dysregulation, polyendocrinopathy, enteropathy, X-linked: forkhead box protein 3 mutations and lack of regulatory T-cells. J Allergy Clin Immunol. 2007 Oct;120(4):744-750

7. Cohen AC, Nadeau KC, Tu W, et al: Cutting edge: Decreased accumulation and regulatory function of CD4+ CD25 (high) T cells in human STAT5b deficiency. J Immunol. 2006 Sep 1;177(5):2770-2774

8. Beyer M, Kochanek M, Giese T, et al: In vivo peripheral expansion of naive CD4+CD25high FOXP3 + regulatory T cells in patients with multiple myeloma. Blood. 2006 May 15;107(10):3940-3949

9. Carmichael KF, Abayomi A: Analysis of diurnal variation of lymphocyte subsets in healthy subjects and its implication in HIV monitoring and treatment. 15th Intl Conference on AIDS, Bangkok, Thailand, 2004, Abstract B11052

10. Dimitrov S, Benedict C, Heutling D, Westermann J, Born J, Lange T: Cortisol and epinephrine control opposing circadian rhythms in T cell subsets. Blood. 2009 May 21;113(21):5134-5143

11. Dimitrov S, Lange T, Nohroudi K, Born J: Number and function of circulating human antigen presenting cells regulated by sleep. Sleep. 2007 Apr;30(4):401-411

12. Kronfol Z, Nair M, Zhang Q, Hill EE, Brown MB: Circadian immune measures in healthy volunteers: relationship to hypothalamic-pituitary-adrenal axis hormones and sympathetic neurotransmitters. Psychosom Med. 1997 Jan-Feb;59(1):42-50

13. Malone JL, Simms TE, Gray GC, Wagner KF, Burge JR, Burke DS: Sources of variability in repeated T-helper lymphocyte counts from human immunodeficiency virus type 1-infected patients: total lymphocyte count fluctuations and diurnal cycle are important. J Acquir Immune Defic Syndr (1988). 1990;3(2):144-151

14. Paglieroni TG, Holland PV: Circannual variation in lymphocyte subsets, revisited. Transfusion. 1994 Jun;34(6):512-516

15. Gambineri E, Ciullini Mannurita S, Hagin D, et al: Clinical, immunological, and molecular heterogeneity of 173 patients with the phenotype of immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome. Front Immunol. 2018 Nov 1;9:2411

16. Park JH, Lee KH, Jeon B, et al: Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome: A systematic review. Autoimmun Rev. 2020 Jun;19(6):102526

17. Delmonte OM, Fleisher TA: Flow cytometry: Surface markers and beyond. J Allergy Clin Immunol. 2019 Feb;143(2):528-537

18. Knight V, Heimall JR, Chong H, et al: A toolkit and framework for optimal laboratory evaluation of individuals with suspected primary immunodeficiency. J Allergy Clin Immunol Pract. 2021 Sep;9(9):3293-3307.e6

Method Description
Describes how the test is performed and provides a method-specific reference

EDTA-anticoagulated blood is incubated with antibodies to various T-cell markers (ie, CD4, CD127, CD45RO, CD45RA, and CD25). After red blood cell lysis, the sample is washed to remove any unbound antibodies prior to analysis. The assay uses 2 antibody tubes for data acquisition, but analysis is performed as a single panel. Each Treg subset is expressed as a percentage of total CD4+ T cells. The regulatory T-cell panel is linked to the TCD4 test (TCD4 / CD4 Count for Immune Monitoring, Blood) within the experiment and, therefore, the CD3, CD4, and CD8 T-cell reference ranges are provided within the TCD4 assay. The regulatory T cell results are interpreted using a reference range derived from data of normal healthy adult and pediatric donors. Isotype controls are used in each assay to measure background fluorescence of the samples. A normal, healthy control is also included in each experiment to ensure the optimal performance of the assay.(Unpublished Mayo information)

PDF Report
Indicates whether the report includes an additional document with charts, images or other enriched information

No

Day(s) Performed
Outlines the days the test is performed. This field reflects the day that the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time before the test is performed. Some tests are listed as continuously performed, which means that assays are performed multiple times during the day.

Monday through Friday

Report Available
The interval of time (receipt of sample at Mayo Clinic Laboratories to results available) taking into account standard setup days and weekends. The first day is the time that it typically takes for a result to be available. The last day is the time it might take, accounting for any necessary repeated testing.

3 to 4 days

Specimen Retention Time
Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded

4 days

Performing Laboratory Location
Indicates the location of the laboratory that performs the test

Rochester

Fees
Several factors determine the fee charged to perform a test. Contact your U.S. or International Regional Manager for information about establishing a fee schedule or to learn more about resources to optimize test selection.

  • Authorized users can sign in to Test Prices for detailed fee information.
  • Clients without access to Test Prices can contact Customer Service 24 hours a day, seven days a week.
  • Prospective clients should contact their Regional Manager. For assistance, contact Customer Service.

Test Classification
Provides information regarding the medical device classification for laboratory test kits and reagents. Tests may be classified as cleared or approved by the US Food and Drug Administration (FDA) and used per manufacturer instructions, or as products that do not undergo full FDA review and approval, and are then labeled as an Analyte Specific Reagent (ASR) product.

This test was developed using an analyte specific reagent. Its performance characteristics were 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
Provides guidance in determining the appropriate Current Procedural Terminology (CPT) code(s) information for each test or profile. The listed CPT codes reflect Mayo Clinic Laboratories interpretation of CPT coding requirements. It is the responsibility of each laboratory to determine correct CPT codes to use for billing.

CPT codes are provided by the performing laboratory.

86359

86361

LOINC® Information
Provides guidance in determining the Logical Observation Identifiers Names and Codes (LOINC) values for the order and results codes of this test. LOINC values are provided by the performing laboratory.

Test Id Test Order Name Order LOINC Value
TREGS T Cell Subsets, Regulatory (Tregs) 90413-6
Result Id Test Result Name Result LOINC Value
Applies only to results expressed in units of measure originally reported by the performing laboratory. These values do not apply to results that are converted to other units of measure.
29149 CD4+CD25+CD127loCD45RA+ Naive Tregs 89315-6
29148 CD4+CD25+CD127loCD45RO+ (Nat Tregs) 89316-4
29147 Activated CD4+ T cells (4+CD25+) 26982-9
29145 % N. Naive Tregs 89319-8
29144 % Natural Tregs 89320-6
29143 % Activated CD4+ T cells (4+CD25+) 13332-2
29146 % CD4+CD25-CD127+ (Tr1/Th3) 89318-0
29150 CD4+CD25-CD127+ (Tr1/Th3) 89317-2
29177 Interpretation 69052-9
609282 CD4 (T Cells) 24467-3

Test Setup Resources

Setup Files
Test setup information contains test file definition details to support order and result interfacing between Mayo Clinic Laboratories and your Laboratory Information System.

Excel | Pdf

Sample Reports
Normal and Abnormal sample reports are provided as references for report appearance.

Normal Reports | Abnormal Reports

SI Sample Reports
International System (SI) of Unit reports are provided for a limited number of tests. These reports are intended for international account use and are only available through MayoLINK accounts that have been defined to receive them.

SI Normal Reports | SI Abnormal Reports