Apoptosis Measurement via Tali Assay

Apoptosis was measured using the Tali™ apoptosis assay kit and propidium iodide (Invitrogen™, Life Technologies, Milan, Italy) as previously reported [24 (link)]. Concisely, on the first day of the assay 1.5 × 10⁵ cells were seeded in a 6-well plate and left to adhere for 16–18 h. The day after seeding, the cells were subjected to UV radiation, with different SPF10-enriched filter combinations, as previously described. At the end of the treatment cells were centrifuged (1500 rpm for 5 min), and resuspended in 100 μL of annexin binding buffer (ABB). After that, 5 μL of Annexin V Alexa Fluor^® 488 was added, mixed well, and the solution was incubated in the dark at room temperature for 20 min. Then cells were centrifuged at 1500 rpm, resuspended in 100 μL of ABB, and 1 μL of propidium iodide was added, mixed well, and incubated in the dark at room temperature for 5 min. Samples were analyzed using the Tali^® Image-Based cytometer and the percentage of apoptotic nuclei, dead cells, and live cells was determined on the basis of the corresponding fluorescence histogram compared with an untreated control. The results were expressed as fold increase compared with the control.