Flow cytometric cytotoxicity assay with GFP gene modified target cells

Abstract

The cytotoxicity assays have been one of the essential tools in studies of effector functions of the immune system. In order to avoid use of radioactive materials many groups have developed alternative ways to quantitate cell mediated lysis. These methods include release of target cell cytoplasmic enzymes (1), dye inclusion or vital dye uptake reduction (2-4). In most of the assays target cells have to be labeled prior analysis which albeit easy to perform is unnecessarily time consuming Here we describe the measurement by flow cytometry killing activities of human NK cells against K562 cells stably transduced to express enhanced green fluorescent protein (EGFP). After short incubation with effector cells killed EGFP positive K562 cells can be later easily identified by propidium iodide (PI) staining. Since EGFP is expressed in every target cell it requires no further cell labelling, cell purification, or extra washing steps thus, it is more time efficient than current assay methods.