Cloning of in vivo-derived thioguanine-resistant human B cells

In vivo-derived thioguanine-resistant (TGr) B cells have been cloned from peripheral blood mononuclear cells (PBMC) of 4 healthy adults. This was done by using Epstein-Barr (EB) virus transformation of B cells enriched from a large number of PBMC obtained with a blood cell separator. The cloned TGr B cells lacked hypoxanthine guanine phosphoribosyltransferase (HPRT) enzyme activity. The frequency of in vivo TGr B cells was estimated to be 8.6-13.1 X 10(-6) for the 4 individuals by comparing the cloning efficiency of non-selected cells and TG-selected cells. This frequency is somewhat higher but comparable to the in vivo frequency of TGr T cells. Because the cloned TGr B cells can be easily expanded in vitro, this procedure provides a large amount of material for the precise characterization of in vivo mutations in humans.