Microstructural Analysis of Gdf5 Knockout Bone

P12 control and Gdf5 KO tibias were dissected, fixated overnight in 4% PFA in PBS, and dehydrated to 100% ethanol (dehydration sequence: 25%, 50%, 70%, and twice in 100% for 1 h each). According to ref. ^{97 (link)}, samples were soaked in 2% iodine in 100% ethanol solution (Sigma) for 48 h at 4 °C. Tissue was washed in 100% ethanol twice for 30 minutes each prior to scanning. Samples were then mounted in 100% ethanol and scanned by MicroXCT-400 (Xradia) at 30 kV and 4.5 W with the Macro-70 lens.

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Rubin S., Agrawal A., Stegmaier J., Krief S., Felsenthal N., Svorai J., Addadi Y., Villoutreix P., Stern T, & Zelzer E. (2021). Application of 3D MAPs pipeline identifies the morphological sequence chondrocytes undergo and the regulatory role of GDF5 in this process. Nature Communications, 12, 5363.

Publication 2021

ethanol solution

Dehydration Ethanol Gdf5 Iodine Lens Tibias Tissue

Corresponding Organization : Laboratoire d’Informatique et Systèmes

Other organizations : Weizmann Institute of Science, RWTH Aachen University

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Variable analysis

independent variables

P12 control
Gdf5 KO

dependent variables

Tibial morphology

control variables

Fixation in 4% PFA in PBS
Dehydration sequence: 25%, 50%, 70%, and twice in 100% ethanol for 1 h each
Soaking in 2% iodine in 100% ethanol solution for 48 h at 4 °C
Washing in 100% ethanol twice for 30 minutes each
Mounting in 100% ethanol
Scanning by MicroXCT-400 at 30 kV and 4.5 W with the Macro-70 lens

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