Immobilized Tetraselmis sp. for ease of TEM processing and for ultrastructure research

Abstract

We report an experimental procedure that results in the immobilization of the motile Tetraselmis in an alginate bead, thereby providing clonal populations for ultrastructure research and greatly facilitating transmission electron microscope (TEM) studies. The Prasinophyte Tetraselmis CS317, which is a potential candidate species for aquaculture, was used in our study. The cells were immobilized in Ca-alginate beads and allowed to grow within the beads for 2 weeks. Each cell in the bead divided repeatedly, resulting in a dense clonal population which could be easily distinguished under a compound microscope. Portions of the Tetraselmis-alginate beads containing the clonal populations were then used for TEM processing for a fine structure study without the need for centrifugation. The normal TEM processing of microalgae by repeated centrifugations during processing or a centrifugation-agar embedding mixture is very time consuming and unreliable due to the nature of the agar and, in the case of the motile Tetraselmis, the force of centrifugation required to sediment the cells. Our results revealed that the alginate did not interfere with the fixation, embedding and sectioning, and the cells appeared to possess all of the structural characteristics of Tetraselmis cells, including the flagella apparatus. We conclude that immobilized Tetraselmis in alginate provides a simple experimental system for ultrastructural research.