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After seeding NSCs within the self-assembled scaffolds as previously described (cell seeding section), cells were cultured for 7 and 14 days in NSC basal medium.
In every analyzed section, cells were quantified in each of the four different regions individually.
As demonstrated in a preceding section, cells transfected with GFP-NACC1-WT contained 10 20 NACC1-NB of various sizes per nucleus.
The percentage of GFP-expressing cells was obtained by dividing the total number of GFP cells per section by the total number of cells in each section; cells were defined by the nucleus label DAPI.
As described in the method section, cells plated at 1 × 104 in soft agar in steroid-depleted medium were treated with AD, GP88 or E2 in the absence of letrozole.
The following features of each experimental time point were quantified cells with elongated shape, cells with pyknotic nuclei, cells on the surface of the section, cells extruded from the section and cells trapped in gelfoam.
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For the results reported in Section "Cell-edge performance", B e corresponds to 60% of the total bandwidth.
For this assay, cells were cultured as described above (section cell culture) for a period of 1, 3, and 7 days (n = 5/time point).
Finally viability was measured using the MTT assay (see section: Cell viability assay).
As described in the Methods section, cell growth medium was exchanged by virus maintenance medium (VVM) at time of infection.
On each section, cell nuclei were stained with Hoechst 33342 (1:500; Sigma-Aldrich) to merge the images exactly.
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