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. 2016 Nov 3;6:36418. doi: 10.1038/srep36418

Figure 2. Localization of ATTO565-HA, ATTO565-sHA1 and FITC-Hep in hBMSC and impact on FN matrix assembly.

Figure 2

hBMSC were cultured on TCPS (control, CTRL, (A). At day 8 after seeding the cells were washed, fixed and stained with mouse anti-human FN-IgG/AlexaFluor488-goat anti-mouse IgG (green fluorescence). The nuclei were visualized by DAPI staining (blue fluorescence). hBMSC were treated from day 1 until day 8 with 200 μg ATTO565-HA/mL (red fluorescence) (B) or 200 μg ATTO565-sHA1/mL (C) and stained at day 8 as described before. hBMSC were treated from day 1 until day 4 (D) with 200 μg ATTO565-sHA1/mL, fixed at day 8 and stained as described before. At day 8, samples, which were treated permanently with ATTO565-sHA1, were decellularized as described and the remaining ECM was fixed and stained for FN (E). hBMSC were treated from day 7 until day 8 (F) with 200 μg ATTO565-sHA1/mL and stained at day 8 as described before. hBMSC were treated with 200 μg ATTO565-sHA1/mL, 5 μg AlexaFluor488-FN/mL and 45 μg plasma-FN/mL. After 48 h cells were fixed and nuclei were stained with DAPI (G). hBMSC were treated with 200 μg FITC-Hep/mL (green fluorescence, labeled in red). At day 8 after seeding the cells were washed, fixed and stained with mouse anti-human FN IgG/AlexaFluor568-goat anti-mouse IgG (red fluorescence, labeled in green) and DAPI (blue) (H). Scale bars A-D and G-H 50 μm, E-F 20 μm.