Harry s hematoxylin

After incubation for 6 min in Harry’s hematoxylin (Carl Roth GmbH and Co.KG), the prepared IVD sections were rinsed in running tap water and counterstained with eosin (Carl Roth GmbH and Co.KG) for 4 min.

The cell sheets were fixed for 15 min with 4% paraformaldehyde (PFA) and placed inside HistoGel (ThermoFisher Scientific Inc., Germany) before being embedded in paraffin. Sections of 7 µm were prepared. The sections were 10 min deparaffinized in xylol (Carl Roth GmbH and Ko.KG) and rehydrated with a descending ethanol row (ETOH, 99.8%, 96%, 80%, 70%) (Carl Roth GmbH and Ko.KG).
For hematoxylin-eosin (HE) staining, sections of sheets were incubated for 6 min in Harry’s hematoxylin (Carl Roth GmbH and Ko.KG), before being rinsed in running tap water and counterstained for 4 min in eosin (Carl Roth GmbH and Ko.KG).
For the alcian blue (AB) stain, sections were incubated for 3 min in 1% acetic acid. Then they were incubated for 30 min in 1% AB staining solution (Carl Roth GmbH and Ko.KG). After rinsing in 3% acetic acid and followed by a washing step in distilled water lasting 2 min, cell nuclei were counterstained with nuclear fast red aluminum sulphate solution (Carl Roth GmbH and Ko.KG) for 5 min. HE- and AB-stained sections were covered with Entellan (Merck KGaA). Images were taken using a light microscope (DM1000 LED, Leica Microsystems GmbH).