live/dead assay kit (calcein am 0.5 μ l/ml and ethidium homodimer-1 (ethd-1, 2 μ l/ml)  (Thermo Fisher)

Journal: BMC Immunology

Article Title: Perforin enhances the granulysin-induced lysis of Listeria innocua in human dendritic cells

doi: 10.1186/1471-2172-8-14

Figure Lengend Snippet: Sublytic perforin does not result in stable pores but induces a transient Ca 2+ flux . L. innocua -challenged DC or non challenged DC were incubated with various concentrations of perforin for 45 minutes at 37°C in the presence of 2 μM EthD-2 in RPMI. After fixation, the percentage of cells that had incorporated the dye was determined by counting three independent experiments by confocal microscopy. As a quality control in parallel experiments, an equal amount of red blood cells were perforin treated for 45 minutes at 37°C before assessing hemolysis.

Article Snippet: For the detection of plasma membrane pores Listeria -challenged DC were incubated with perforin at the indicated concentration for 45 minutes at 37°C in medium containing 2 μM ethidium homodimer-2 (EthD-2, Invitrogen).

Techniques: Incubation, Confocal Microscopy