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. 2017 Mar 3;7:43669. doi: 10.1038/srep43669

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Copyright © 2017, The Author(s)

This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

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(a) Hela-GFP and Hela cells were mixed 1:1, treated with 0.1 μM rosuvastatin or 20 μM betulin for 8days, fixed in 3.7% paraformaldeide and incubated with the primary antibody anti-neuroserpin (Ab32901, Abcam) coniugated PLA probes (1:50), overnight at 4 °C. The presence of aggregates are visualized using Duolink in Situ staining according to the manufacturer’s instructions. Nuclei are stained with DAPI. Images are acquired by Leica SP2 laser scanning confocal microscope. Each red spot represents a neuroserpin aggregate. The number of spots per cell for each condition in the images showed in this panel is quantified as described in the methods section. (b) Untreated or treated cells with 0.1 μM rosuvastatin or 20 μM betulin (8 days) were fixed with 3.7% paraformaldeide and then incubated with TNM-AMCA (1 μM) to stain cholesterol for 1 h at room temperature. Images were acquired by Leica SP2 laser scanning confocal microscope.