Quantifying Cartilage Proteoglycan Content

Proteoglycan content in cartilaginous tissue was measured by dimethylmethylene Blue (DMMB) assay, as described before (Melnik et al., 2019 (link)), and the values were normalized to DNA amount in lysed cells measured with Quant-iT PicoGreen dsDNA kit (Invitrogen, Eugene, United States). For this, 20 μl of the digested pellet sample were mixed with 80 μl TE buffer (200 mM Tris HCl, 20 mM EDTA) and PicoGreen solution, and fluorescence in samples was measured at 485/535 nm.

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Melnik S., Hofmann N., Gabler J., Hecht N, & Richter W. (2021). MiR-181a Targets RSPO2 and Regulates Bone Morphogenetic Protein – WNT Signaling Crosstalk During Chondrogenic Differentiation of Mesenchymal Stromal Cells. Frontiers in Cell and Developmental Biology, 9, 747057.

Publication 2021

Quant-iT PicoGreen dsDNA kit

Assay Buffer Cartilaginous tissue Cells Dimethylmethylene blue Dsdna Edta Fluorescence Picogreen Proteoglycan Tris

Corresponding Organization : Heidelberg University

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Variable analysis

independent variables

Proteoglycan content in cartilaginous tissue

dependent variables

DNA amount in lysed cells

control variables

DMMB assay as described in Melnik et al., 2019
Quant-iT PicoGreen dsDNA kit (Invitrogen, Eugene, United States)
Mixing 20 μl of the digested pellet sample with 80 μl TE buffer (200 mM Tris HCl, 20 mM EDTA) and PicoGreen solution
Measuring fluorescence in samples at 485/535 nm

controls

No positive or negative controls were explicitly mentioned in the provided information.

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