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Short answer
Terminal deoxynucleotidyl transferase (TdT) is a DNA polymerase that extends the 3′ end of a DNA strand one nucleotide at a time, without a template. In the immune system it adds random nucleotides during V(D)J recombination. In the lab it is the enzyme behind TUNEL labelling and 3′ tailing. In pathology, TdT staining marks immature lymphoid cells. This is general information, not medical advice; a TdT result on a patient report is interpreted by the treating clinicians.
This guide connects those uses and tabulates, from open-access reviews, the metals TdT uses and where it is reported positive or negative.
Definition and scope
The NC-IUBMB enzyme list gives TdT the accepted name DNA nucleotidylexotransferase, EC 2.7.7.31. It describes the reaction as template-independent addition of a 2′-deoxyribonucleoside 5′-triphosphate to a DNA chain, and notes that the enzyme cannot start a chain de novo. Motea and Berdis (2010) describe TdT as the one polymerase that can add nucleotides using only a single-stranded DNA initiator.
What is the function of transferase enzymes?
A transferase moves a chemical group from one molecule to another. The IUBMB classification makes transferases class EC 2 and divides them by the group moved: one-carbon groups (EC 2.1, which includes the methyltransferases covered in our DNA methyltransferase guide), glycosyl groups (EC 2.4) and phosphorus-containing groups (EC 2.7). TdT sits in EC 2.7.7, the nucleotidyltransferases: it transfers a nucleotide from a dNTP onto a DNA 3′ end.
How TdT works and how it is used
What are the four types of dNTPs?
The four deoxynucleoside triphosphates are dATP, dCTP, dGTP and dTTP (deoxyadenosine, deoxycytidine, deoxyguanosine and deoxythymidine triphosphate). Motea and Berdis define a classical DNA polymerase as an enzyme that adds mononucleotides to a primer, guided by a template, in the presence of the four dNTPs. TdT uses the same chemistry with no template. See dGTP and its role in DNA synthesis and how nucleotides build DNA.
Which metals TdT uses
Like all DNA polymerases, TdT needs a divalent metal ion, and it can use several. The table summarises Motea and Berdis (2010); the ranking applies to extending a p(dA)n primer with dATP.
| Metal | Reported effect on TdT |
|---|---|
| Mg²⁺ | Fastest extension of the p(dA)n primer with dATP; favours dGTP and dATP |
| Zn²⁺ | Second fastest; micromolar Zn²⁺ added to Mg²⁺ reactions stimulates the rate |
| Co²⁺ | Third; increases efficiency with the pyrimidines dCTP and dTTP |
| Mn²⁺ | Slowest of the four; supports the reaction less efficiently than Mg²⁺ |
3′ tailing and 3′ end labelling
Because TdT extends 3′-OH ends, 3′ tailing means adding a run of nucleotides to a DNA 3′ end, and 3′ end labelling means adding a labelled nucleotide there. Motea and Berdis report that TdT accepts nucleotide analogues, tolerates bulky modifications to the base, and can label the 3′ termini of synthetic oligonucleotides with a radioactive nucleotide or a variety of fluorescent probes. An analogue with no 3′-OH, such as cordycepin, ends extension when incorporated. A different non-templated addition, the single A that Taq polymerase puts on PCR products, is described in our TA cloning guide.
TUNEL
The same property drives TUNEL: TdT incorporates biotinylated dUMP onto the 3′ ends of single-stranded DNA at DNA breaks, and the biotin is visualised with fluorescently labelled avidin or streptavidin. The method is widely used to detect apoptosis. Label chemistries and controls are compared in our TUNEL assay guide. In TUNEL the enzyme is a reagent; in a TdT stain (below) the enzyme is what is being detected.
TdT in V(D)J recombination
After the RAG proteins cut the DNA and Artemis:DNA-PKcs opens the hairpins at the coding ends, TdT randomly adds nucleotides to the available 3′-OH ends, which increases variability in the non-templated (N) regions of the recombined antigen receptor genes. Motea and Berdis give the average N segment as only 2 to 5 bp per coding joint, with an in vivo bias towards dGMP and dCMP, and place TdT in the primary lymphoid tissues, the thymus and bone marrow. This is why a polymerase also works as a marker of immature lymphoid cells.
Types and related concepts
Which cancers are TdT positive?
TdT is characteristic of lymphoblastic leukemia and lymphoma, the neoplasms of immature B or T lymphoid precursors. Motea and Berdis report variable TdT expression in about 90% of patients with acute lymphoblastic leukemia.
| Cell type or disease | TdT as reported | Source |
|---|---|---|
| B-lymphoblastic leukemia/lymphoma | Part of the typical immature phenotype, including the pro-B stage; cases with KMT2A rearrangement are typically TdT-negative | Dorfman 2025 |
| T-lymphoblastic leukemia/lymphoma | Typically positive, seen as nuclear staining on immunohistochemistry; one illustrated TdT-negative case is called not typical | Danielson 2026 |
| NK-lymphoblastic leukemia/lymphoma | 50% of six cases in one study were positive | Dorfman 2025 |
| Acute myeloid leukemia | Typically negative for TdT in the pathology review; the enzyme review reports TdT overexpression in about 20% of AML | Danielson 2026; Motea and Berdis 2010 |
| T-prolymphocytic leukemia | No TdT expression | Danielson 2026 |
| Burkitt lymphoma | Negative for TdT | Danielson 2026 |
| Thymoma (immature T-cell lymphoid component) | Can show TdT positivity and resemble T-lymphoblastic lymphoma; separated on histology | Danielson 2026 |
| Normal early B-cell precursors (hematogones) | Type I are TdT-positive; TdT falls as they mature | Dorfman 2025 |
What are the functions of DNA polymerase 1, 2, and 3?
The numbering is the Escherichia coli one, as described by Makiela-Dzbenska and colleagues (2009). TdT is not one of the three; it belongs to the pol X family with pol μ and pol λ, whose roles in V(D)J recombination are distinct and non-overlapping in vivo (Motea and Berdis).
- Pol III does most chromosomal replication. Its holoenzyme synthesises both strands; the α subunit polymerises and the ε subunit proofreads with a 3’→5′ exonuclease.
- Pol II (polB) is a high-fidelity B-family enzyme with 3’→5′ exonuclease activity. It is proposed to stand in for Pol III when that enzyme struggles to extend a mismatched primer end.
- Pol I (polA) was the first polymerase discovered and is the most abundant (about 400 molecules per cell). Its 5’→3′ exonuclease removes the RNA primers of Okazaki fragments, and it fills gaps during repair.
Where it matters for labs and clinics
A TdT stain or TdT marker result comes from detecting the protein in cells, by immunohistochemistry (nuclear staining) or flow cytometry. Danielson and colleagues note that blast lineage cannot be reliably determined by morphology alone, so flow cytometry and immunohistochemistry are essential. Dorfman describes the diagnostic approach as combining morphology, immunophenotype (usually by flow cytometry) and genetic analysis, so TdT is read alongside other lineage markers. For staining controls, see our immunohistochemistry guide.
Common misunderstandings
- “TdT positive means cancer.” Normal early B-cell precursors and the immature lymphoid component of thymoma can be TdT-positive.
- “TdT and TUNEL are the same test.” TUNEL uses the enzyme to label DNA breaks. A TdT stain detects the enzyme protein in the cell.
- “TdT is the only polymerase that adds untemplated nucleotides.” Others do so on blunt duplex DNA; TdT is the one that works from a single-stranded initiator.
FAQ
What does TdT positive mean?
In the pathology reviews cited here, TdT staining indicates an immature lymphoid cell. It is interpreted with other markers, not alone.
Which cancers are TdT positive?
Mainly B- and T-lymphoblastic leukemia/lymphoma; see the table for variations, including KMT2A-rearranged B-lymphoblastic leukemia and AML.
What are the four types of dNTPs?
dATP, dCTP, dGTP and dTTP.
What is the function of transferase enzymes?
They transfer a chemical group between molecules; TdT transfers a nucleotide onto a DNA 3′ end.
What are the functions of DNA polymerase 1, 2, and 3?
In E. coli, Pol III replicates the chromosome, Pol I removes RNA primers and fills repair gaps, and Pol II is a high-fidelity polymerase proposed to back up Pol III.
Sources
- Motea EA, Berdis AJ. Terminal deoxynucleotidyl transferase: the story of a misguided DNA polymerase. Biochim Biophys Acta, 2010;1804(5):1151-1166, checked 2026-11-07.
- Danielson D, Lagerstrom IT, Rogers M, Simon K, et al. T-lymphoblastic leukemia/lymphoma: a comprehensive review of pathology, molecular features, and differential diagnosis. J Pathol Transl Med, 2026;60(5):502-515, checked 2026-11-07.
- Dorfman DM. The flow cytometric evaluation of B- and T-lymphoblastic leukemia/lymphoma. Cancers (Basel), 2025;17(7):1111, checked 2026-11-07.
- Makiela-Dzbenska K, Jaszczur M, Banach-Orlowska M, Jonczyk P, et al. Role of Escherichia coli DNA polymerase I in chromosomal DNA replication fidelity. Mol Microbiol, 2009;74(5):1114-1127, checked 2026-11-07.
- NC-IUBMB Enzyme Nomenclature, EC 2.7.7.31, DNA nucleotidylexotransferase and EC 2 Transferases, checked 2026-11-07.
