The CD300c (CMRF-35A) and CD300a (CMRF-35H) molecules are leukocyte surface proteins that are part of a larger family of immunoregulatory molecules encoded by a gene complex on human chromosome 17. The CMRF-35 monoclonal antibody binds to an epitope common to both molecules, expressed on most human leukocyte populations, apart from B lymphocytes and a subpopulation of CD4(+) and CD8(+) T lymphocytes. We describe the CMRF-35(pos) and CMRF-35(-) fractions of CD4(+) T lymphocytes. The CMRF-35(pos) fraction can further be divided into CMRF-35(++) and CMRF-35(+)CD4(+) T lymphocyte subpopulations. Resting peripheral CD4(+) T lymphocytes express CD300a mRNA and very low amounts of CD300c. Activation results in an initial decrease in CD300a gene expression before an increase in both CD300a and CD300c gene expression. The up-regulated expression of these genes was associated with increased CMRF-35 binding to activated T lymphocytes. The CMRF-35(-) fraction of CD4(+) T lymphocytes proliferated to a greater extent than the CMRF-35(pos) fraction, in response to mitogens or allogeneic antigen. The poor proliferation of the CMRF-35(pos) CD4(+) in response to mitogens was explained by increased apoptosis within this subpopulation. The recall antigen, tetanus toxoid, stimulated the CMRF-35(++)CD4(+)CD45RO(+) but not the CMRF-35(-)CD4(+)CD45RO(+) subpopulation. Resting CMRF-35(++) CD4(+) lymphocytes express low levels of IFN-gamma mRNA. Within 18 h following in vitro activation, CMRF-35(++) CD4(+) lymphocytes express more IFN-gamma mRNA and protein compared with the CMRF-35(-)CD4(+) lymphocytes, however, after 24 h, both the CMRF-35(+) and CMRF-35(-)CD4(+) T lymphocytes were able to produce IFN-gamma. The CMRF-35(++)CD4(+) T lymphocyte population contains the Th(1) memory effector cells.