Micron hm355s

Mice were anaesthetized via injection with a xylazine (20 mg/kg)/ketamine (200 mg/kg) solution and sacrificed by cervical dislocation after blood puncture via the retro-orbital vein and cardiac flushing with PBS. Thyroid lobes were excised and either snap-frozen for RNA analysis or cut into 1 mm³ pieces for tissue dissociation and flow cytometry analysis. For histological or immunolabelling analysis on paraffin sections, thyroid lobes were extracted with the trachea, fixed in 4% paraformaldehyde at 4 °C overnight, and embedded in paraffin using a Tissue-Tek VIP-6 (Sakura). Sections of 6 µm were obtained with the microtome Micron HM355S (Thermo Scientific, Waltham, MA, USA). For immunohistofluorescence on frozen sections, thyroid lobes were collected and fixed in 4% paraformaldehyde at 4 °C overnight, followed by an overnight immersion in PBS/20% sucrose solution. Finally, tissues were embedded in PBS/15% sucrose/7.5% gelatin solution and stored at −80 °C. Sections of 7 µm were obtained with a cryostat (Thermo Scientific, Cryostar NX70). Frozen human tissue samples (sections or tissue pieces) were obtained from the Saint-Luc Hospital’s Biobank, Brussels. The study followed the Declaration of Helsinki and was approved by the local ethics committee of the UCLouvain, Brussels, Belgium (2017/25OCT/495 and 2017/04OCT/466).

Mice were sacrificed by cervical dislocation under anesthesia by 150 µL of a xylazine (20 mg/kg)/ketamine (200 mg/kg) solution and after cardiac puncture. Thyroid lobes were excised and either snap-frozen for RNA analysis or cut in 1 mm³ pieces for tissue dissociation and EV isolation. For histological and immunohistochemical analyses, thyroid lobes were extracted together with trachea, fixed in 4% paraformaldehyde and embedded in paraffin using a Tissue-Tek VIP-6 (Sakura, Torrance, CA, USA). Sections of 6 µm were obtained with the microtome Micron HM355S (ThermoScientific, Waltham, MA, USA). After paraffin removal and rehydration, histological sections were stained with hematoxylin and eosin, slides were mounted in Dako aqueous Medium (Agilent Technologies, Santa Clara, CA, USA) and scanned with the panoramic P250 digital slide scanner (3DHistech, Budapest, Hungary), as described in [28 (link)].

The caudal half of E12.5, E14.5 and E18.5 CD1 embryos were fixed in 4% formaldehyde for 1 h and embedded in paraffin using a Tissue-Tek VIP-6 (Sakura). Sections of 7 µm were obtained with the microtome Micron HM355S (ThermoScientific). After paraffin removal, slides were treated similarly as gelatin sections.