EXPERIMENTAL FIGURE 23-22 Chromium (⁵¹Cr) release assay allows the direct demonstration of the cytotoxicity and specificity of cytotoxic T cells in a heterogeneous population of cells. (a) A suspension of spleen cells containing cytotoxic (killer) T cells is prepared from mice that have been exposed to a particular virus (e.g., influenza virus) and have cleared the infection. Target cells obtained from mice of the same strain are infected with the identical virus or left uninfected. After infection, cellular proteins are labeled nonspecifically by incubation of the target-cell suspension with ⁵¹Cr. When the radiolabeled target cells are incubated with the suspension of cytotoxic T cells, the killing of infected target cells results in release of the ⁵¹Cr-labeled proteins. Uninfected target cells are not killed and retain their radioactive contents. Lysis of cells by cytotoxic T cells can therefore be readily detected and quantitated by measuring the radioactivity released into the supernatant. (b) Cytotoxic T cells (CTLs) harvested from mice that have been infected with virus X can be tested against various target cells to determine the specificity of CTL-mediated killing. CTLs capable of lysing virus X–infected target cells 1 cannot kill uninfected cells 2 or cells infected with a different virus, Y 3. When these CTLs are tested on virus X–infected targets from a strain of mice that carries an altogether different MHC type (strain b), again no killing is observed 4. Cytotoxic T-cell activity is thus virus specific and restricted by the MHC.