THE HUMAN T-CELL RECEPTOR V-BETA REPERTOIRE OF NORMAL PERIPHERAL-BLOOD LYMPHOCYTES BEFORE AND AFTER MITOGEN STIMULATION

Mitogen stimulation of T cells in vitro has been employed in the analy sis of the T cell antigen receptor (TCR) repertoire and as a method of generating T cell lines and clones. It has been suspected for some ti me that mitogen stimulation may bias the repertoire. We have addressed this problem employing a semi-quantitative technique utilizing the po lymerase chain reaction (PCR) and flow cytometry. Using this PCR metho d and a panel of primers to 22 Vbeta subgroups, the Vbeta repertoire o f both unstimulated and phytohaemagglutinin (PHA)-stimulated periphera l T cells from eight healthy individuals was investigated. The samples were also analysed by flow cytometry using anti-Vbeta2, Vbeta5 and Vb eta8 MoAbs. A significant increase in the expression of Vbeta6, Vbeta7 .2 and Vbeta10. 1 was found in all eight samples of PHA-stimulated T c ells compared with unstimulated T cells using the PCR method. In contr ast, no differences were found between unstimulated and PHA-stimulated T cells by flow cytometry. These results question the validity of usi ng mitogen-stimulated T cells to investigate TCR gene usage.