Detection and determination of autophagy in SK-N-BE2 and IMR-32 cells after treatment with 200 nM rapamycin for 6, 12, and 24 h.

<p>Here, cells without any treatment were used as the untreated control (CTL). All experiments were performed in triplicates. (a) Detection of autophagic populations. Cells were treated with rapamycin and then stained with acridin orange (AO) for flow cytometric detection of the acidic vesicular organelles (AVO) in autophagic cells. (b) Determination of percentages of autophagic populations after AO staining. Significant difference between CTL and a treatment time was indicated by *<i>P</i><0.05 whereas significant difference between a treatment time and another treatment time was indicated by #<i>P</i><0.05. (c) Western blotting to show the protein levels of LC3 I and LC3 II forms in both cell lines. Expression of β-actin was used as a loading control in Western blotting.