Cytotoxic activity of splenocytes from tumor-bearing A/J mice after ch14.18–IL-2 fusion protein therapy. (A) Experimental metastasis was induced by intravenous injection of 1 × 10⁶ NXS2 cells followed by six daily intravenous administrations of either 10 μg ch14.18–IL-2, an equivalent mixture of ch14.18 antibody and rhIL-2, or PBS. Twenty-four hours after completion of the treatment, the cytotoxic activity of splenocytes was tested in a 4-hour chromium release assay against YAC1 (closed symbols) and P815 (open symbols) target cells. (B) Splenocytes of immunocompetent (closed symbols) and in vivo–depleted mice (open symbols) were tested for lysis against YAC1 cells in a 4-hour chromium release assay 24 hours after completion of the treatment with PBS or 10 μg ch14.18–IL-2 fusion protein (daily, ×6, starting 24 hours after induction of experimental metastases). Immunodepletion was started at day −3 and −1 before tumor cell inoculation followed by a once-weekly schedule with intraperitoneal injection of either 350 μg anti-CD8 antibody or 100 μL anti-asialo GM1 antiserum, respectively. (C) Splenocytes of tumor-bearing mice treated with 10 μg ch14.18–IL-2 fusion protein (daily, ×6, starting 24 hours after induction of experimental metastases) were tested for lysis of NXS2 target cells in a 4-hour chromium release assay. Splenocytes were used at an effector-to-target-cell ratio of 100:1 in the presence of (a) PBS; (b) 30,000 IU/mL rhIL-2; (c) 10 μg/mL ch14.18; (d) 30,000 IU/mL rhIL-2+10 μg/mL ch14.18; and (e) 10 μg/mL ch14.18–IL-2. The lysis of NK-cell–enriched (f) and pure CD8⁺ T-cell (g) subfractions was compared with that of whole splenocytes (e) in the presence of 10 μg/mL ch14.18–IL-2.