Pacific oyster Crassostrea gigas is one major species for aquaculture, and the development of breeding programs and the need for preservation of wild stock genetic resources prompted the need for larvae cryopreservation. The objective of the present study was to choose the most reliable protocol from several existing publications, to test its biological and practical limitations, and to adapt it to hatchery conditions. The selected protocol was characterized by a very slow freezing rate without seeding, and by the use of ethylene glycol and sucrose as cryoprotectant. The best survivals after thawing and rearing up to 48 h post fertilization (hpf) were obtained with larvae that were frozen at late trochophore (20 hpf) and early-D (24 hpf) stages. Increasing the larvae concentration in the straws and using high throughput straw filling and freezing devices did not alter the cryopreservation outcome. The whole procedure was applied to cryopreservation in a commercial hatchery (Satmar, France), and the thawed larvae yielded 9.4 ± 4.5% survivals at 12 days post fertilization. The overall success was dampened by some variability in the larvae survival that is likely due to the physiological status of the larvae. In all, the proposed procedure is robust and reliable and can be used for cryobanking of oyster genetic resources.