Tracking living decapod larvae: mass staining of eggs with neutral red prior to hatching

Abstract

Mass staining of decapod females carrying eggs, with subsequent identification of hatched larvae in the environment, is a research tool with great potential for field ecologists wishing to track the movements of larvae. For this to be achieved, however, numerous requirements must be met. These include adequate dye solubility, short staining time, dye penetration through different tissues, dye retention within the organism, absence of toxic and behavioral effects, low visibility to predators of stained larvae, no loss of staining owing to preservatives and low cost. The dye, neutral red, appears to meet most of these requirements. This dye was used in aliquots of 0.7 g/770 ml seawater applied to the females of Norway lobster (Nephrops norvegicus) and European lobster (Homarus gammarus) for 10 min. This procedure stained lobster eggs and embryos so that hatched larvae could be distinguished easily by fluorescence microscopy from larvae that hatched from unstained eggs. Stained larvae that were preserved in 4% formaldehyde in seawater were still stained after 1 year. Larvae should not come in contact with ethanol, because it extracts the dye rapidly.