Efficacy of glyoxal acid-free (GAF®) as a non-toxic fixative for fish gonads, gametes and larvae: widespread implications for laboratory studies, field sampling and applied aquaculture

Traditional fixatives such as buffered formaldehyde (BF) and paraformaldehyde (PFA) are widely used but raise health concerns due to their toxicity. Their use is increasingly subject to regulatory restrictions, calling for the development of non-toxic alternatives. Glyoxal acid-free (GAF®) has emerged as a potential substitute. Aims and methods: This study evaluated GAF® for preserving gonadal tissues, oocyte, sperm, and larvae of the European seabass (Dicentrarchus labrax), comparing both its standard and an optimized, marine-adapted formulation to conventional fixatives or fresh samples. Key results: The standard GAF® led to tissue shrinkage and poor gamete preservation, while the optimized version effectively preserved oocyte, sperm, gonadal tissue, and delicate larval structures. Although 10% formalin still outperformed GAF® for certain larval tissues, extended fixation in optimized GAF® improved results. Conclusions: Overall, optimized GAF® represents a promising non-toxic alternative for histological applications in marine fish, though further refinement is needed for certain tissues and for the use in the oocyte cytoplasmic clarification applied in aquaculture broodstock selection.