The world population is rapidly growing and is predicted to grow to 10 billion by 2050, as estimated by the food and Agriculture Organization (FAO), requiring a 70% rise in total food production and 100% increase in meat production to satisfy nutritional requirements and global demand. Thus, developing novel technologies to produce food products is needed.
This study aimed to optimize primary cell line development from adult Eastern Oyster, Crassostrea virginica, as a model bivalve by applying different tissue decontamination, cell dissociation, and culture conditions. Oyster adductor (OAD) cells were obtained via tissue explant, mechanical and enzymatic digestion. Culture vessels were coated with surface proteins such as fibronectin, laminin, Matrigel, and poly-d-lysine to promote cell attachment. The tissue decontamination with Penicillin-Streptomycin (100 µg/mL), Amphotericin B (0.25 µg/ml), and algaecide solution (0.03%) were effective in controlling contaminations. OAD cells grew best at lower nutrient levels in the one-to-one ratio of Lebovitz L-15 media and artificial seawater. Lower fetal bovine serum levels, 1-5%, provided a high number of cell attachments and consistent growth in combination with 1% adult oyster whole-body or larvae extract. The tissue explant method resulted in optimal cell dissociation compared to other methods, and proceeding cultures had attached cells surviving for up to 10 days. All the plate coatings promoted cell attachment. However, fibronectin provided optimal cell attachment of OAD cells. Fibroblast-like, neuron-like, epithelial-like, and rounded cells were observed. Fluorescence cell staining confirmed the presence of cytoskeleton and nuclei in the OAD cell cultures. These advances in primary cell culture methods of OAD cells may be beneficial for establishing mollusk cell lines for cultivated seafood production.