Abstract: The invention is intended to further improve the operability, economic efficiency and safety in the preparation of antigen-specific CTLs. The invention provides a preparation kit used for a method for preparing antigen-specific cytotoxic T lymphocytes, the method comprising: a first step for inducing antigen-specific cytotoxic T lymphocytes, wherein the components of the first step include a culture medium contained in an injection vessel, a hermetically scaled culture vessel, and the like; a second step for preparing an activated T cell for antigen presentation, wherein the components of the second step include a culture medium contained in an injection vessel, a hermetically sealed culture vessel, and the like.; and a third step for proliferating antigen-specific cytotoxic T lymphocytes, wherein the components of the third step include a culture medium contained in an injection vessel, a hermetically sealed separation vessel, a hermetically sealed culture vessel, and the like.