Cryopreservation of Balanus amphitritenauplii

Abstract

The possibility of cryopreservingBalanus amphitrite(Cirripedia; thoracica) nauplii is explored. The effects on the postthaw survival of a range of cryobiological variables are reported; three different cryoprotectants (ethylene glycol, EG; dimethyl sulfoxide, Me₂SO; and glycerol, GL), the influence of the final temperature (−20, −30, −40, and −50°C), and the effect of transferring to liquid nitrogen (−196°C) were studied. The larvae showed increasing susceptibility to cryoprotectant concentration. Equilibration of larvae for 15 min at 5°C with each of the cryoprotectants revealed that GL was the least toxic. EG at 3 to 4Mdid not cause marked damage as the equilibration period was extended up to 2 h. This was not true with Me₂SO and GL. The postthaw survival rate decreased as the final temperature was reduced, and none of the larvae revived after thawing from −50°C. The postthaw survival with 2 mol/liter EG as the cryoprotectant ranged from 86% after cooling to −20°C to 56% after cooling to −40°C. The larvae that were transferred to liquid nitrogen from different temperatures revealed that the greatest survival (36%) was achieved when the larvae were transferred from −40 to −196°C. When the larvae were reared after cooling to −40°C and then transferring to liquid nitrogen, the rate of subsequent metamorphosis was 30% in the case of EG at 3M. Me₂SO and GL gave results that were inferior in comparison to those obtained with EG.