Figure 4

Extent of apoptosis determined by subdiploid DNA and nuclear localization of AIF. Logarithmically growing A549 cells were seeded, incubated with CuO NP, CuO MP or CuCl₂ for 24 h, fixed and (A) stained with antibodies against AIF for fluorescent microscopy (evaluation of at least 50 cells per condition) or (B) stained with DAPI for flow cytometry analysis of sub-G1 peak (evaluation of 10 000 cells per condition) as described in Materials and methods. The data represent mean values of at least 3 independent experiments + SD. Statistically significant different from control: *p < 0.05, **p < 0.01, ***p < 0.001 as determined by unifactorial analysis of variance (ANOVA) and Dunnett’s T3 test. 1 μg/mL are equal to 0.2 μg/cm² and 50 μg/mL CuO are equal to 630 μM Cu²⁺ in case of complete dissolution. n.d.: not determined.