The effects of triiodothyronine (T3), thymulin, and recombinant chicken interferon-γ (ChIFN-γ) on natural killer (NK) cell cytotoxicity was investigated using the euthyroid control K and the T3-deficient sex-linked dwarf (SLD) chicken strains. Factorial design experiments were used to investigate the effects of T3 treatments where animals of both strains received either 0 or 0.1 ppm T3 supplementation to the standard chick starter diet. The ChIFN-γ treatments were administered in vitro by incubation with effector cells overnight prior to the addition of the RP9 lymphoblastoma target cell line. All cytotoxicity assays were run at 50:1 and 25:1 effector/target (E/T) ratios. Treatments were begun at hatching and continued through 7 weeks. NK cells for these assays were enriched by separation of splenocytes over ficoll. Splenocyte preparations from untreated K strain consistently had significantly higher NK-mediated cytolysis than did samples from the untreated SLD at both E/T ratios. T3 treatment alone had no effect on NK activity in cell preparations from the K strain but did significantly enhance that activity in the T3-deficient SLD whereas IFN treatment alone enhanced NK activity in both strains. The combined T3 and IFN treatments resulted in a greater enhancement of NK cytolytic activity in both strains than any separate treatment and resulted in an elimination of differences in NK cell responsiveness between the K and SLD strains. Similar results were obtained when NK cell cultures were incubated in vitro with thymulin prior to assessing cytotoxicity. In vitro thymulin treatments alone significantly enhanced cytolytic activity for NK cells for both K and SLD strains. The greatest effect of in vitro thymulin exposure was to increase the responsiveness to NK cells to ChIFN-γ stimulation.