Methods of sample preparation of radula epithelial tissue in chitons (Mollusca: Polyplacophora)

A glutaraldehyde fixative developed for preserving the radula superior epithelium of the adult chiton Acanthopleura hirtosa (Blainville, 1825), was used in conjunction with conventional and microwave-assisted sample processing to produce high quality tissue preservation for light and electron microscopy. In addition, high-pressure freezing (HPF) and cryo-substitution were used to fix the radula tissue of juvenile specimens. Microwave- assisted fixation was preferred to conventional bench-top techniques due to the superior preservation of fine cell structure together with reduced processing dines and chemical exposure. Although restricted to very small (<200 im) samples, the quality of juvenile radulae processed by HPF was excellent. The improvements in tissue preservation using microwave and cryo-preservation techniques are therefore critical for obtaining accurate ultrastructural information on the radula in marine molluscs. In particular, these findings highlight additional processing options available for the study of cellular structures in biomineralizing tissues.