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Blood, 1 July 2001, Vol. 98, No. 1, pp. 165-173
IMMUNOBIOLOGY
Molecular and flow cytometric analysis of the V repertoire for
clonality assessment in mature TCR T-cell proliferations
Anton W. Langerak,
René van den Beemd,
Ingrid L. M. Wolvers-Tettero,
Patrick P. C. Boor,
Ellen G. van Lochem,
Herbert Hooijkaas, and
Jacques J. M. van
Dongen
From the Department of Immunology, University Hospital
Rotterdam/Erasmus University Rotterdam, The Netherlands.
Clonality assessment through Southern blot (SB) analysis of
TCRB genes or polymerase chain reaction (PCR) analysis of
TCRG genes is important for diagnosing suspect mature
T-cell proliferations. Clonality assessment through reverse
transcription (RT)-PCR analysis of V -C transcripts and flow
cytometry with a V antibody panel covering more than 65% of V
domains was validated using 28 SB-defined clonal T-cell receptor
(TCR) + T-ALL samples and T-cell lines. Next, the
diagnostic applicability of the V RT-PCR and flow
cytometric clonality assays was studied in 47 mature T-cell
proliferations. Clonal V-C RT-PCR products were detected in all
47 samples, whereas single V domain usage was found in 31 (66%) of
47 patients. The suspect leukemic cell populations in the other 16 patients showed a complete lack of V monoclonal antibody reactivity
that was confirmed by molecular data showing the usage of V
gene segments not covered by the applied V monoclonal antibodies.
Nevertheless, this could be considered indirect evidence for the
"clonal" character of these cells. Remarkably, RT-PCR revealed an
oligoclonal pattern in addition to dominant V-C products and
single V domain expression in many T-LGL proliferations, providing
further evidence for the hypothesis raised earlier that T-LGL derive
from polyclonal and oligoclonal proliferations of antigen-activated
cytotoxic T cells. It is concluded that molecular V analysis
serves to assess clonality in suspect T-cell proliferations. However,
the faster and cheaper V antibody studies can be used as a powerful
screening method for the detection of single V domain expression,
followed by molecular studies in patients with more than 20% single
V domain expression or large suspect T-cell populations (more than
50%-60%) without V reactivity.
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