Dynamic changes of accumulated T cell clonotypes during antigenic stimulation in vivo and in vitro

It has not been known how T cell clones respond to antigenic stimulation. We applied reverse transcription polymerase chain reaction and subsequent single-strand conformation polymorphism (SSCP) analysis to monitor the kinetics of accumulated T cell clonotypes with respect to the TCR beta chain. Before exposure to an antigen, peripheral blood lymphocytes from healthy individuals exhibited smears with SSCP, indicating a heterogeneous T cell population. However, some bands were demonstrated within the smears, each corresponding to a distinct clonal accumulation. The majority of the accumulated clones were CD8+ and were stably detected over 1 year in the same individual. Next it was demonstrated that a number of clonal accumulations appeared in a diverse population after an acute infection in vivo or in cell culture with an antigen in vitro. The accumulations in vivo were demonstrated in both the CD4+ and CD8+ subsets, the latter having the longer duration of response after stimuli. On the other hand, in vitro culture with purified protein derivative induced expansions of a number of CD4+ clones throughout the culture, while CD8+ clones were shown to be induced later. There was no preferential usage of particular V beta genes in these clonotypes. Stimuli by more simple peptides could also induce clonal expansions of T cells. Therefore, with our novel experimental system, we could monitor the kinetics of T cell clonotypes induced by an antigen. This method may be applied to analyses of T cell clonality in many kinds of immune responses.