Flow cytometric analysis of the V? repertoire in healthy controls

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Abstract
Analysis of the T-cell receptor (TCR)-Vβ repertoire has been used for studying selective T-cell responses in autoimmune disease, alloreactivity in transplantation, and protective immunity against microbial and tumor antigens. For the interpretation of these studies, we need information about the Vβ repertoire usage in healthy individuals. We analyzed blood T-lymphocyte (sub)populations of 36 healthy controls (age range: from neonates to 86 years) with a carefully selected most complete panel of 22 Vβ monoclonal antibodies, which together recognized 70–75% of all blood TCRαβ+ T lymphocytes. Subsequently, we developed a six-tube test kit with selected Vβ antibody combinations for easy and rapid detection of single (“clonal”) Vβ domain usage in large T-cell expansions. The mean values of the Vβ repertoire usage were stable during aging in blood TCRαβ+ T lymphocytes as well as in the CD4+ and CD8+ T-cell subsets, although the standard deviations increased in the elderly. The increased standard deviations were caused by the occurrence of oligoclonal T-cell expansions in the elderly, mainly consisting of CD8+ T lymphocytes. The 15 detected T-cell expansions did not reach 40% of total TCRαβ+ T lymphocytes and represented less than 0.4 × 109 cells per liter in our study. Vβ usage of the CD4+ and CD8+ subsets was comparable for most tested Vβ domains, but significant differences (P < 0.01) between the two subsets were found for Vβ2, Vβ5.1, Vβ6.7, Vβ9.1, and Vβ22 (higher in CD4+), as well as for Vβ1, Vβ7.1, Vβ14, and Vβ23 (higher in CD8+). Finally, single Vβ domain expression in large T-cell expansions can indeed be detected by the six-tube test kit. The results of our study can now be used as reference values in studies on distortions of the Vβ repertoire in disease states. The six-tube test kit can be used for detection of single Vβ domain expression in large T-cell expansions (>2.0 × 109/l), which are clinically suspicious of T-cell leukemia. Cytometry 40:336–345, 2000

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