Use of monoclonal antibodies to study alloreactivity and T cell development in Xenopus

Abstract

This study investigated T-cell surface antigenic changes in the spleen of Xenopus following in vivo and in vitro allergenic challenge, using novel mouse monoclonal antibodies (mAb's) against all T cells (CDS), the putative ɑβ-T cell receptor (TCR), the putative γδ-TCR, the CD8 receptor and against major histocompatibility complex (MHC) class II proteins. Flow cytometric analyses of splenocytes from skin allografted intact, 5-day thymectomised (Tx) and skin allotolerant animals were examined. There was an increase in the number of cells expressing γδ TCR's in those froges which rejected test allografts. The Tx Xenopus laevis and allotolerant LG3 animals that tolerated grafts showed no increase in the 76 TCR. Twenty-four days post-grafting X. laevis, most of the γδ TCR positive cells surprisingly co-expressed the ɑβ TCR. Following mixed lymphocyte reactions (MLR) for 9-14 days, of previously grafted X. laevis and LG15 splenocytes, there was an increase in the number of T cells with a significant increase γδ-TCR positive cells. Stimulation indices of MLR's were increased by pre-culturing the stimulator cells with concanavalin A (ConA). This increase was apparently not due to changes in cytokine production or cell surface antigen expression, of the stimulated cells. Little increased MHC class II expression was seen, in flow cytometric analyses. The reasons for elevated MLR with ConA-activated stimulators therefore remain to be assessed. The in vitro response of Xenopus splenocytes to the superantigen, staphylococcal enterotoxin B (SEB), was tested. The response was weakly significant but was significantly less than that seen in mice.