Provided are methods of isolating and proliferating antigen-specific CD8+ T cells using anti-4-1BB antibodies. The methods of isolating and proliferating CD8+ T cells may yield cells at a higher recovery rate than a conventional isolation method, and the isolation method using humanized anti-4-1BB antibodies may also yield cells at a high recovery rate. Further, in cell culture, cells may be grown at a high proliferation rate. The antigen-specific CD8+ T cells yielded according to the isolation and proliferation methods may be used to treat cancer without any side-effects.