A reliable anaesthesia and sampling protocol for Pacific oysters will enable experiments to be conducted without sacrificing animals and will facilitate successive sampling of individuals for gametogenesis studies. As no such techniques were available for Crassostrea gigas, the present study aimed to define suitable anaesthetic conditions for use with this species. Three groups of ten oysters (mean weight ± SD, 32.1 ± 9.0 g) were anaesthetised in 5 L containers. Among different chemicals: benzocaine, eugenol and three different types of magnesium chloride (a laboratory one – Flucka® – and two designed for agriculture – DEUSA International® and Dead Sea Works®) and concentrations tested, one laboratory (concentration: 72 g L−1) and one agricultural (Dead Sea Works® 50 g L−1) type of magnesium chloride were the most effective, respectively inducing anaesthesia in 73 ± 3% and 80 ± 3$% after three hours. Lower oyster weight and a two day period of starving prior to treatment significantly increased the number of anaesthetised animals. Using this protocol, losses of 0 to 10% of oysters were observed one week after anaesthesia. Increasing anaesthesia duration from 3 to 16 h resulted in a significant increase in the number of anaesthetised oysters (from 50 ± 10 to 97 ± 7%) but no increase in mortality (7 ± 11%). On the other hand, reducing water temperature from 19.5 °C to 15.3 °C, resulted in a significant decrease in anaesthesia efficacy. A reliable anaesthesia protocol was developed: 100% of Pacific oysters are anaesthetised using 50 g L−1MgCl2 Dead Sea Work®for a 16 h duration. This long duration facilitates tissue sampling and does not increase the working time needed. This protocol was validated by monthly anaesthesia and gonad sampling during a three month period with the loss of only a single oyster. It can thus be used for routine applications.