Technique for Revealing the Three-Dimensional Architecture of Whole Preserved Spiculated Invertebrates

Abstract

Procedures for revealing the three-dimensional arrangement of calcareous sclerites, spicules, or ossicles embedded within connective tissue in formalin fixed invertebrates are described. Spicules are stained with alizarin red S following maceration of preserved animals or colonies with either trypsin or KOH solutions. Connective tissue is stained with alcian blue in different samples prior to maceration. Stained animals or colonies are cleared in glycerin. This method for revealing spicular structure and arrangement and the gross morphology of connective tissues offers several advantages over either scanning electron microscopy or reconstruction from serial sections.