The quality of oyster spermatozoa has become an issue for modern aquaculture. However, reliable protocols used to assess gamete quality are lacking. The aim of this work is to define a standardised experimental fertilisation protocol for Pacific oyster oocytes. Six experiments have been carried out in this study. The optimal conditions for Pacific oyster experimental fertilisation have been defined: (1) oocytes can be conserved in seawater for at least 4 h before fertilisation. (2) oocyte concentration: 100-1000 ml(-1) seawater, (3) fertilisation volume: 10-100 nil, (4) spermatozoa: oocyte ratio: 400 to maintain discriminating conditions, (5) gamete contact time: longer than 10 mill and (6) fertilisation and incubation temperatures: both 19 degrees C. The effect of individual male significantly influenced the fertilisation success. The results of this study will be exploited for spermatozoal quality analysis in combination with other techniques. (C) 2008 Elsevier B.V. All rights reserved.