2.7. Cell size and shape measurement
Neuronal microtubules were identified by β-III-Tubulin immunostaining, as described above and in triplicate cultures for each sample. A total of 100 cells were assessed per subject from 15 random microscopy fields. The area and perimeter occupied by microtubules was defined and measured with the ImageJ software (USA). The formula used to assess roundness was: (perimeter2)/(4*pi*area). A roundness value of 1.0 indicates a perfect circle, and higher values indicate oblong and non-circular objects (DeCoster et al., 2010).