Leonardo D. Mendoza-González Lucia Suárez López ^* CARMEN G. PANIAGUA CHAVEZ ^*

• Center for Scientific Research and Higher Education in Ensenada (CICESE), Ensenada, Mexico

The cryopreservation of cell lines such as primordial germ cells and germ cells is a promising strategy to conserve and reconstitute endangered or commercially important species in aquaculture. In Mexico, the northwest region concentrates the country's most significant fishing and aquaculture production. However, most of the species used in capture fishing are overexploited. Despite this, protocols for cryopreservation of germ cells are non-existent. Therefore, this work aimed to establish a protocol of isolation, identification, and cryopreservation of germ cells in two species, Totoaba (Totoaba macdonaldi) and Yellowtail Amberjack (Seriola lalandi). Three concentrations of trypsin (0.25%, 0.3%, and 0.5%) were tested for gonadal dissociation. The 0.3% trypsin concentration was the best because it presented the most significant number of viable cells, with 14.35×10 5 for Totoaba and 2.96×10 5 for Yellowtail Amberjack. The immunohistochemistry identification of germ cells in both species was positive for vasa, with 33.30% for Totoaba and 34.20% for Yellowtail Amberjack. The cryoprotectant used was ethylene glycol (1.5M or 2M). The ideal temperature for cryopreservation of gonadal tissue was different for each species, -1°C/min for Totoaba and -5°C/min for Yellowtail amberjack with 58.42% and 63.48% viable cells after thawing respectively, being ethylene glycol 1.5M the best for both species. The noncontrolled rate was the most effective technique to freeze cell suspension, with 4.20 ± 1.09×10 5 /mL viable cells for Totoaba and 7.31 ± 2.25×10 5 /mL for Yellowtail Amberjack. In conclusion, the results of isolation, identification, and cryopreservation protocols for germ cells in Totoaba and Yellowtail amberjack obtained in this work is the first report for fish species from northwest Mexico, opening the door for the generation of cryobanking of germ cells.Finally, this work would help conserve endangered species and be an alternative to conserving species of commercial importance in aquaculture.