Decalcification and Tissue Observation of Coral

For tissue section observations, the fixed coral fragments were decalcified according to Kopp’s method (50 (link)). In detail, the fixed coral fragments were decalcified at 4°C in Sorensen-sucrose phosphate buffer containing 0.5 M EDTA. The decalcification buffer was renewed daily until being completely demineralized. Rinsed coral tissue samples were dissected and post-fixed in 1% OsO₄ in Sorensen-sucrose phosphate buffer for 1 hour at room temperature, and then dehydrated in ethanol and embedded in Spurr resin. Sections were cut with a Diamote 35° diamond (Ultracut microtome). Tissue sections were stained with HE (hematoxylin-eosin staining) and observed with a light microscope LEICA DMRB equipped with a LEICI DC300F camera (Leica, France).