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Immunophenotyping by Flow Cytometry: Double Negative T Cells (DNT) Test

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Immunophenotyping by Flow Cytometry: Double Negative T Cells (DNT) Test

Short Name: DNT Test by Flow Cytometry

Also known as: Double Negative T Cell Test, DNT Cell Analysis, CD4-CD8- T Cell Test, TCRαβ+ CD4-CD8- T Cell Immunophenotyping, ALPS Screen – DNT Cells

Immunophenotyping by Flow Cytometry: Double Negative T Cells (DNT) Test test available at DNA Labs India for ₹5,500. Uses Flow Cytometry, Multi-color Immunophenotyping on Whole Blood samples. Results in Sample Daily by 9 am; Report Next Working Day. Free home collection in 300+ cities across India.

Flow Cytometry🏠 Home Collection

✍️ Written By

🩺 Medically Reviewed By

Dr Pasupathy Arumugam

Dr Pasupathy Arumugam

Consultant Pathologist · Reg: 21521

Last reviewed: September 7, 2026

Overview

The purpose of the DNT test is to identify and quantify Double Negative T cells (CD3+ TCRαβ+ CD4- CD8-) in peripheral blood using flow cytometry. This test is primarily used as a screening tool for Autoimmune Lymphoproliferative Syndrome (ALPS) and to support the evaluation of other autoimmune and inflammatory conditions. Elevated DNT cells may indicate impaired lymphocyte apoptosis and are an important biomarker in the diagnostic workup of lymphoproliferative disorders.

Test Code
1090
Price
₹5,500
Sample Type
Whole Blood
Result Time
Sample Daily by 9 am; Report Next Working Day
Fasting Required
No
Method
Flow Cytometry, Multi-color Immunophenotyping
Step 1

Sample Collection

No fasting is required. Provide a brief clinical history and relevant family history on the test request form. Inform the healthcare provider about any current medications, especially immunosuppressive drugs.

Method: Venipuncture

Step 2

Laboratory Analysis

A blood sample will be collected from a vein in your arm (venipuncture). Two separate tubes will be drawn: one Lavender Top (EDTA) tube and one Green Top (Sodium Heparin) tube, each containing a minimum of 2 mL of whole blood.

Step 3

Report Delivery

Apply gentle pressure to the puncture site with a cotton ball or gauze for a few minutes. Normal activities can be resumed immediately. Mild bruising at the site is normal and resolves on its own.

Timeline: Sample Daily by 9 am; Report Next Working Day

Patient Instructions

1
Before the Test:No fasting is required. Provide a brief clinical history, family history, and current medication list on the test request form. Ensure samples are collected and shipped at room temperature (18–22°C) on the same day.
2
During the Test:Two blood samples will be collected via venipuncture — one in an EDTA (Lavender Top) tube and one in a Sodium Heparin (Green Top) tube. The procedure typically takes 5–10 minutes and involves minimal discomfort.
3
After the Test:Apply gentle pressure to the venipuncture site. Normal activities can be resumed immediately. Mild bruising may occur and resolves on its own. Reports are typically available the next working day after sample receipt.

About This Test

Who Should Get This Test

The purpose of the DNT test is to identify and quantify Double Negative T cells (CD3+ TCRαβ+ CD4- CD8-) in peripheral blood using flow cytometry. This test is primarily used as a screening tool for Autoimmune Lymphoproliferative Syndrome (ALPS) and to support the evaluation of other autoimmune and inflammatory conditions. Elevated DNT cells may indicate impaired lymphocyte apoptosis and are an important biomarker in the diagnostic workup of lymphoproliferative disorders.

How to Prepare

  • Collect 3 mL (minimum 2 mL) whole blood in 1 Lavender Top (EDTA) tube
  • Collect 3 mL (minimum 2 mL) whole blood in 1 Green Top (Sodium Heparin) tube
  • Ship samples immediately at 18–22°C (room temperature)
  • DO NOT refrigerate or freeze the samples
  • Specify time of collection, date, and clinical details on the test request form
  • Avoid hemolysis during blood draw

Doctor's Notes

Reviewed by — MBBS, MD (Pathology) · Reg. No. 21521

"As an immunologist, I recommend the DNT test for patients presenting with unexplained lymphoproliferation, multilineage cytopenias, or autoimmune manifestations suggestive of Autoimmune Lymphoproliferative Syndrome (ALPS). The identification of elevated TCRαβ+ CD4- CD8- double negative T cells — typically exceeding 1.5% of total lymphocytes or 2.5% of CD3+ T cells — raises suspicion for ALPS and warrants further genetic evaluation. However, mild elevations may also be observed in other autoimmune conditions such as SLE and rheumatoid arthritis. This test should always be interpreted in the context of clinical findings, family history, and complementary laboratory investigations including serum vitamin B12, soluble FAS ligand levels, and FAS/FASL/CASP10 genetic testing."

Last medically reviewed: September 7, 2026

Test Parameters & Specifications

Sample TypeWhole Blood
Sample Volume3 mL (2 mL min.) in EDTA tube AND 3 mL (2 mL min.) in Sodium Heparin tube
Container1 Lavender Top (EDTA) tube AND 1 Green Top (Sodium Heparin) tube
Collection MethodVenipuncture

Sample Stability

Room Temperature (18–22°C)
Refrigerated (2–8°C)
Frozen
Sample Rejection Criteria:
  • Refrigerated or frozen samples
  • Clotted samples
  • Hemolyzed specimens
  • Insufficient sample volume (less than 2 mL per tube)
  • Samples received after 24 hours of collection at room temperature
  • Samples without clinical details or time of collection on request form
  • Incorrect tube type used

Understanding Your Results

The DNT test measures the percentage of Double Negative T cells (CD3+ TCRαβ+ CD4- CD8-) in peripheral blood using multi-color flow cytometry. Results must be interpreted alongside the patient's clinical presentation, family history, and supplementary investigations including genetic testing.
📊

Normal finding. Consistent with healthy individuals. No further DNT-specific evaluation typically required.

📊

Borderline elevation. Clinical correlation recommended. Repeat testing and additional investigations (serum vitamin B12, soluble FAS ligand) may be advised.

📊

Elevated DNT cells. Suggestive of ALPS or other autoimmune/lymphoproliferative conditions. Genetic testing for FAS, FASL, or CASP10 gene mutations is recommended for confirmation.

📊

Strongly suggestive of ALPS. Urgent referral to a clinical immunologist and comprehensive genetic evaluation is warranted.

⚠️ When to Consult a Doctor:

Consult your doctor if you experience persistent unexplained lymphadenopathy (swollen lymph nodes), splenomegaly (enlarged spleen), recurrent autoimmune cytopenias (low blood cell counts), or symptoms suggestive of autoimmune conditions such as joint pain, fatigue, skin rashes, or unexplained fevers. Early evaluation is particularly important if there is a family history of autoimmune lymphoproliferative disorders or ALPS.

Limitations

  • ⚠Not a confirmatory diagnostic test; abnormal results require correlation with clinical history and genetic testing
  • ⚠Elevated DNT cells may be observed in conditions other than ALPS, including SLE and other autoimmune diseases
  • ⚠Results may be affected by improper sample handling — samples must be shipped at 18–22°C and must not be refrigerated or frozen
  • ⚠Immunosuppressive therapy may reduce DNT cell counts, potentially leading to false-negative results
  • ⚠Technical variability between laboratories may affect reference range interpretation

Risks & Considerations

  • ●Minor bruising or discomfort at the venipuncture site
  • ●Very rare risk of infection at the puncture site
  • ●Lightheadedness or dizziness during blood draw in sensitive individuals

Interfering Factors

  • ●Recent blood transfusion may alter lymphocyte population ratios
  • ●Immunosuppressive medications (e.g., corticosteroids, methotrexate, rituximab) may affect DNT cell counts
  • ●Sample stored at incorrect temperature (refrigerated or frozen) will yield unreliable results
  • ●Clotted or hemolyzed specimens are not suitable for flow cytometry analysis
  • ●Ongoing chemotherapy or biologic therapy may suppress or alter T cell subsets

Compare With Similar Tests

TestImmunophenotyping by Flow Cytometry: Double Negative T Cells (DNT) Test
ComparisonImmunophenotyping by Flow Cytometry: Double Negative T Cells (DNT) TestThe lymphocyte subset panel provides a broad overview of immune cell populations. The DNT test is a more focused analysis specifically targeting the CD4-CD8- double negative T cell subset for ALPS screening.FAS gene mutation analysis is a confirmatory genetic test for ALPS. The DNT test serves as an initial screening tool; if elevated, FAS mutation analysis is recommended to confirm the diagnosis.sFASL measures a soluble apoptotic marker in serum. When combined with the DNT test, it improves the diagnostic sensitivity for ALPS.

Frequently Asked Questions

What is a Double Negative T Cell (DNT) test?
The DNT test is a specialized flow cytometry-based immunophenotyping test that identifies and quantifies Double Negative T cells (CD3+ TCR??+ CD4- CD8-) in peripheral blood. These are T cells that express the alpha-beta T cell receptor but lack both CD4 and CD8 co-receptors. The test is primarily used as a screening tool for Autoimmune Lymphoproliferative Syndrome (ALPS).
What is the purpose of the DNT test?
The purpose of the DNT test is to screen for Autoimmune Lymphoproliferative Syndrome (ALPS) and to support the evaluation of other autoimmune and inflammatory conditions. Elevated DNT cells are a hallmark finding in ALPS and indicate impaired lymphocyte apoptosis.
What sample is required for the DNT test?
Two blood samples are required: 3 mL (minimum 2 mL) of whole blood in a Lavender Top (EDTA) tube and 3 mL (minimum 2 mL) of whole blood in a Green Top (Sodium Heparin) tube. Both samples must be shipped immediately at 18–22°C and must not be refrigerated or frozen.
Is fasting required for the DNT test?
No, fasting is not required for the DNT test. However, you should provide a brief clinical history, family history, and a list of current medications on the test request form.
What conditions are associated with elevated DNT cells?
Elevated DNT cells are most strongly associated with Autoimmune Lymphoproliferative Syndrome (ALPS). They may also be found in other autoimmune and inflammatory conditions such as systemic lupus erythematosus (SLE), rheumatoid arthritis, and multiple sclerosis.
What is Autoimmune Lymphoproliferative Syndrome (ALPS)?
ALPS is a rare genetic disorder caused by defects in the apoptosis (programmed cell death) pathway of lymphocytes. It is commonly associated with mutations in the FAS, FASL, or CASP10 genes. Patients with ALPS typically present with chronic lymphadenopathy, splenomegaly, autoimmune cytopenias, and an elevated population of Double Negative T cells.
How is the DNT test performed?
The DNT test is performed using multi-color flow cytometry. Blood samples are stained with fluorescent antibodies targeting specific cell surface markers (CD3, CD4, CD8, TCR??). The flow cytometer then analyzes thousands of cells per second, identifying and quantifying the DNT cell population based on their unique marker expression pattern.
What is the normal range for DNT cells?
In healthy individuals, DNT cells (CD3+ TCR??+ CD4- CD8-) typically constitute less than 1% of total peripheral blood lymphocytes, or less than 2.5% of CD3+ TCR??+ T cells. Values above these thresholds warrant further clinical evaluation and may indicate ALPS or other autoimmune conditions.
Is the DNT test a confirmatory test for ALPS?
No, the DNT test is a screening test and is not confirmatory on its own. Abnormal results should be correlated with the patient's clinical history and confirmed through genetic testing for relevant gene mutations (FAS, FASL, CASP10) and additional laboratory investigations such as serum soluble FAS ligand levels.
What is the cost of the DNT test at DNA Labs India?
The cost of the Immunophenotyping by Flow Cytometry: Double Negative T Cells (DNT) test at DNA Labs India is INR 5500. This includes free home sample collection across India and report delivery via online portal, email, and WhatsApp.
How long does it take to get DNT test results?
If the sample is received daily by 9 am, the report is typically available on the next working day. Reports are delivered through the online portal, email, and WhatsApp for your convenience.
Can the DNT test be performed for children?
Yes, the DNT test can be performed for patients of all age groups, including children. It is particularly relevant for pediatric patients suspected of having ALPS, as the condition often presents in early childhood. A pediatrician or clinical immunologist may recommend this test based on clinical findings.
Worried about the process? Our certified phlebotomists collect thousands of samples every month across India. The process takes under 5 minutes and is virtually painless. Questions? Message us on WhatsApp — we're here 7 days a week.

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