miRNA library screen in differential trogocytosis of CTL

CTL-mediated killing is often accompanied by the exchange of cell surface material between CTL and target cells, which is know as trogocytosis. We used it as a proxy for identifying the most lethal CTLs. Murine OT1 CTLs were retrovirally transduced with one of five miRNA pools and then incubated with OVA-loaded EL4 target cells with one of three different membrane stains. Two hours later, CTLs with three kinds of fluorescent labels ("best" killer), CTLs without acquired stain ("worst" killer) were sorted and subjected to next generation sequencing to identify miRNAs that were enriched in the trogocytosis-hi or the trogocytosis-lo subsets. In this manner, we were able to identify miRNAs that reproducibly altered cytotoxic ability in vitro.