This invention provides a non-radioactive
assay to monitor and quantify the target-
cell killing activities mediated by cytotoxic T lymphocytes (CTLs). This
assay is predicated on the discovery that
apoptosis pathway activation and, in particular,
caspase activity, provides a measure of cytotoxic
effector cell activity. In one embodiment, measurement of CTL-induced
caspase activation in target cells is achieved through detection of the specific cleavage of fluorogenic
caspase substrates. This
assay reliably detects
antigen-specific CTL killing of target cells, and provides a more sensitive, more informative and
safer alternative to the standard 51Cr-release assay most often used to quantify CTL responses. The assay can be used to study CTL-mediated killing of primary host target cells of different
cell lineages, and enables the study of
antigen-specific cellular immune responses in real time at the single-cell level. As such, the assay can provide a valuable tool for studies of infectious
disease pathogenesis and development of new vaccines and immunotherapies.