Tracking alloreactive cell division in vivo

Background. The study of alloimmune responses has been limited by a lack of assays that can track the behavior of alloreactive lymphocytes in vivo. He re we utilize an experimental system that allows the identification and stu dy of alloreactive CD4(+) lymphocytes responding to major histocompatibilit y antigens in vivo. Methods. Responder mouse lymphocytes were labeled with a fluorescein-based dye, adoptively transferred into irradiated allogeneic stimulator mice, and recovered at serial time points for analysis by flow cytometry. Results. Discrete generations of CD4(+) responder lymphocytes proliferating specifically in response to allogeneic MHC class II were distinguished by fluorescein intensity. Successive division of alloreactive CD4(+) lymphocyt es was traced up to six generations after 60 hr. Conclusions. This experimental system provides information on the division kinetics of alloreactive CD4(+) cells. Other applications include immunophe notyping of alloreactive lymphocyte subsets. Further study of systems such as this will allow the detailed characterization of how alloimmune response s are initiated and proceed in vivo.