In vivo killing of antigen-specific T lymphocytes using K^b-CD28-ζ and K^b-CD28[L→G]-ζ RMTCs. A total of 10⁷ CFSE-labeled OT-1 lymph node cells were adoptively transferred intravenously into SCID mice; 10⁷ Kb-CD28-ζ or K^b-CD28[L→G]-ζ peptide-pulsed or unpulsed RMTCs were then adoptively transferred intravenously at an anatomically separate location. Twenty-four hours after transfer, spleen and mixed lymph nodes (mesenteric, cervical, axillary, inguinal) were isolated and single-cell suspensions prepared, stained with V_α2-specific antibody, and analyzed by flow cytometry. (A) Percentage of transferred CFSE-positive OT-1 cells that are V_α2-positive (ovalbumin-specific) or V_α2-negative (nontarget cells) in the spleens of mice receiving different RMTC treatments is shown. RMTC effector and ovalbumin peptide pulsing is indicated on the abscissa. For each RMTC effector and peptide-pulse combination, data from individual mice are indicated with a unique symbol (♦, ▴, ▪). Mean percentages are indicated by a horizontal bar. (B) As in panel A, but LNs of recipient animals were analyzed. (C) Normalized numbers of target cells in the spleens of treated animals. The ratio of residual transferred (CFSE-positive) RMTC targets (V_α2-positive) to nontargets (V_α2-negative) was calculated to control for the efficiency of adoptive transfer in mice treated with peptide-pulsed or control unpulsed effectors. (D) Analysis of LN cells. Error bars show ± 1 SD. Results are representative of 3 independent experiments.