Effects of cryoprotectant on the embryos of banded coral shrimp (Stenopus hispidus); preliminary studies to establish freezing protocols.

The addition of cryoprotectants is a necessary step in cryopreservation procedures because they can minimize cellular injury during cryopreservation. Toxicity of cryoprotectant depends on the type, concentration, temperature and exposure period. The aim of this study was to investigate the toxicity of cryoprotectant to embryos of banded coral shrimp (Stenopus hispidus) in order to inform the development of a cryopreservation protocol. Three stages of embryonic development (eye-formation, heart beat and pre-hatch stage) embryos were selected and exposed to different concentration of cryoprotectants (0.25M-5M) for an equilibration period of 10, 20 or 30 min at room temperature. Hatching percentage indicated that the toxicity of tested cryoprotectants increased in the order of methanol, ethylene glycol (EG), dimethyl sulphoxide (DMSO), glycerol and dimethylacetamide (DMA). The No Observed Effect Concentrations (NOECs) for eye-formation stage embryos were 1M, 0.25M, 0.25M, 0.25M and 0.25M respectively after 10 min incubation whilst the NOECs for heart beat and pre-hatch stage embryos were 1M, 0.5M, 0.5M, 0.5M and 0.25M respectively. Pre-hatch stage embryos appeared to be more tolerant to cryoprotectant toxicity than eye-formation and heart beat stage embryos.