%0 Figure %A Trajkovic, Katarina %A Valdez, Clarissa %A Ysselstein, Daniel %A Krainc, Dimitri %D 2019 %T Cell density affects experimental outcomes. %U https://plos.figshare.com/articles/figure/Cell_density_affects_experimental_outcomes_/7670003 %R 10.1371/journal.pone.0211727.g006 %2 https://plos.figshare.com/ndownloader/files/14248421 %K Lamin B 1. %K lysosomal enzyme cathepsin D %K proteins HDAC 1 %K cell population density %K autophagic markers p 62 %K plasma membrane proteins Na %K cell culture-based studies %K cell culture data %K cell density %K LC %K FAK %K cell cycle checkpoint regulator p-cdc 2 %K 3II %X

(A) HEK 293FT cells were plated in 6 well-plates according to the following scheme: 5 sets of control cells at a range of densities (100-500K cells per well) and 4 sets of cells at the highest density (500K cells per well) for treatment. All control cells were treated with the vehicle (control), and the remaining 4 sets were treated with 25 μM Ly294002 (Ly), 2.5 μg/μl Brefeldin A (BFA), 100 ng/ml nocodazole (noco) or 800 nM YM-201636 (YM), for the last 20h before lysis. Ctl N, control cells plated at the same number (500K) as the treated cells; Ctl D, control cells with matching cell density at the experimental endpoint. In classical experimental design, Ctl N is used as a control. (B) Cells were imaged before the lysis and matched with one of the controls based on cell density. Scale bar, 100 μm. (C-F) Cell lysates were analyzed by Western blotting using indicated antibodies. Ctl N and Ctl D were simultaneously compared with treated cells. GAPDH was used as a loading control. (G-J) Western blot images were quantified and the values normalized to GAPDH; Bar graph data are mean ± SD. *p<0.05, **p<0.01, ***p<0.001.

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