The elicitation of delayed-type hypersensitivity (DTH) responses in mice is mediated by the sequential activities of two different Ag-specific, Thy-1+ cells. A required early phase of elicitation is due to DTH-initiating Thy-1+ cells that are CD3- and sIg- and produce Ag-specific factors that act like IgE antibodies in that they sensitize the tissues, so that after local challenge with Ag there is release of the vasoactive amine serotonin. Released serotonin locally recruits and activates CD4+ Th-1 classical DTH effector T cells that secrete lymphokines that attract and activate a nonspecific perivascular infiltrate of circulating, bone marrow-derived leukocytes. The current study used isolated subpopulations of DTH-initiating and DTH-effector T cells to determine whether the two phases of the elicitation of DTH were entirely separate. The contact sensitivity model of DTH was used. Early-acting DTH-initiating cells, and late-acting DTH-effector T cells were either from oxazolone (OX)-immune or picryl chloride (PCl)-immune CBA or BALB/c donors and were transferred to CBA or BALB/c recipients. The results showed that DTH-initiation could be mediated by polyclonal DTH-initiating cells that were Ag mismatched or MHC incompatible with late-acting DTH effector T cells. In fact DTH-initiating cells could be both Ag mismatched and MHC incompatible with late-acting T cells. In addition, potential interactions between different cell populations were ruled out by showing that DTH-initiation could be mediated by a DTH-initiating clone that was Ag or MHC mismatched with the late-acting DTH-effector T cells. Thus, the OX-specific BALB/c clone could initiate DTH for PCl-specific CBA cells in CBA recipients if the recipients were challenged with both OX and PCl, but not when they were challenged with OX or PCl alone. We suggest, at least for the elicitation of DTH reactions in mice, that a more comprehensive description of these responses should accommodate the fact that there are early and late phase responses that each begin with Ag specificity and end with non-specific humoral factors. Inasmuch as the two Thy-1+ cells of DTH can be of different Ag specificity, this suggests that some forms of delayed and chronic inflammation, might be initiated by an immediate hypersensitivity-like immune reactivity to one set of Ag, and could be prolonged and perpetuated by delayed reactivity to another set of Ag.