Analysis of TdT expression in cytometry practice

The terminal deoxynucleotidyl transferase (TdT) is a nuclear DNA polymerase that is primarily found in immature lymphoid cells of the B- and T-cell lineages. It´s biological function is to catalyse the addition of N-nucleotides to the V, D, and J exons of the TCR and BCR genes during recombination. This proces induces variability of antigen receptors and plays an important role in the evolution of immune system in vertebrates. TdT is an immaturity-associated marker, the level of which is the highest in the physiological progenitors of B-lymphocytes in the phase of IGH gene rearrangements in early progenitor B cells and gradually decreasing with B cell maturation. In the T-cell line, we can detect TdT in immature thymic cells; during further differentiation, its expression disappears. TdT is located exclusively in the cell nucleus, so for its detection it is necessary to use intracytoplasmic staining after permeabilization of both cellular and nuclear membrane using fixation /permaebilization reagents.

In cytometry practice, the analysis of TdT expression is used in the diagnostics of acute lymphoblastic leukemia, or to distinguish B-Lymphoblastic Leukemia / Lymphoma with Burkitt-like Morphology (TdT +) from Burkitt Lymphoma (TdT-)(1).

In acute myeloid leukemia we can find coexpression of TdT most often in cases with translocation t (8; 21) (q22; q22) (2).

TdT positivity has also been reported in high-grade B-cell lymphoma, with MYC and BCL2 and / or BCL6 rearrangements. For these reasons, it is not recommended to use the TdT marker to determine the lineage of the neoplastic population. (3)

The use of TdT for monitoring of minimal residual disease is also limited because of the possibility of steroid-induced loss of immaturity-associated antigens (CD10, CD34, and TdT) during induction therapy of acute leukemias. (4)

Fig. 1: Flow cytometry intracellular (permeabilization and fixation ED7397) staining pattern of Aute Myeloid Leukemia (AML) patient stained using anti-human TDT (41A) FITC antibody. Red are TDT positive leukocytes. Clinicaly tested by RNDr. Martin Novák, Ph.D.

In our laboratory, we have successfully tested the new EXBIO anti-Hu TdT FITC antibody (Clone 41A), which in combination with the fixation / permeabilizing agent EXCELLYSE XPerm shows excellent results in both peripheral blood and bone marrow samples.
 

Martin Novak, Ph.D.
Laboratory of Flow Cytometry / Department of Hematooncology
Olomouc University Hospital

References:
1 Li Y, Gupta G, Molofsky A, Xie Y, Shihabi N, McCormick J, Jaffe ES. B Lymphoblastic Leukemia/Lymphoma With Burkitt-like Morphology and IGH/MYC Rearrangement: Report of 3 Cases in Adult Patients. Am J Surg Pathol. 2018 Feb;42(2):269-276 2 Hurwitz CA, Raimondi SC, Head D, Krance R, Mirro J Jr, Kalwinsky DK, Ayers GD,Behm FG. Distinctive immunophenotypic features of t(8;21)(q22;q22) acute myeloblastic leukemia in children. Blood. 1992 Dec 15;80(12):3182-8. 3 Swerdlow SH, Campo E, Harris NL, et al. WHO classification of tumors of hematopoietic and lymphoid tissues. Revised 4th edition. Lyon: International Agency for Research on Cancer; 2017 4 Gaipa G, Basso G, Maglia O, et al. Drug-induced immunophenotypic modulation in childhood ALL: implications for minimal residual disease detection. Leukemia. 2005;19(1):49–56.

EXBIO PRODUCTS RELATED TO THIS BLOG:
 

• Anti-Hu TdT FITC antibody, 41A clone, Cat. Number: 1F-868-T100
• EXCELLYSE XPerm ( a set of two solutions intended for red blood cell lysis together with permeabilization of cytoplasmic membranes of leukocytes in the preparation of biological samples from human peripheral blood prior to the flow cytometry analysis), Cat. Number: ED7397
• Mouse IgG2a Isotype Control FITC, Cat. Number: 1F-724-C100

Coming soon product: Anti-Hu TdT PE antibody, 41A clone, Cat. Number: 1P-868-T100