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Fig. 2 | Stem Cell Research & Therapy

Fig. 2

From: Automated digital image quantification of histological staining for the analysis of the trilineage differentiation potential of mesenchymal stem cells

Fig. 2

Validation of the DIA approach. The DIA approach was compared to absorbance measurements and showed an increased signal to noise ratio for both osteogenic and adipogenic differentiation (a). (n = 18 for osteogenic differentiation, n = 9 for adipogenic differentiation, error bars represent SD). Six different cell lines were differentiated into osteoblasts and adipocytes and afterwards analyzed with DIA and absorbance measurements (b). (n = 9 for all cell lines except for F22 n = 3, error bars represent SD). The ranking of all six cell lines showed a signal overestimation for absorbance measurements and a superior sensitivity of the DIA approach (c). Correlation analysis for osteogenesis revealed that DIA does reliably quantify Alizarin Red S. The correlation between absorbance measurements and DIA for adipocytes is less strong (d). The reason for this is a signal overestimation originating from absorbance measurements for weakly differentiated cells (e.g., controls). DIA also allows to quantify histological dyes that cannot be quantified by absorbance (e). Chondrogenic differentiation was quantified with DIA using Alcian Blue 8GX-stained cells (n = 9, error bars represent SD). Chondroblasts that were differentiated in 3D were cut into multiple cross sections, and positive pixels were compared to the total number of pixels of the spheroid (f) leading to a quantitative expression of the chondrogenic differentiation (g)

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