Considering the concerns about the safety of the use of formaldehyde as antifungal agent, the effects of three alternative chemicals (potassium sorbate, copper hydroxide and magnesium chloride) were tested in the artificial incubation of signal crayfish (Pacifastacus leniusculus) eggs. Eight treatments were performed during 15 min every other day: formaldehyde at 3000 ppm (control), potassium sorbate at 5000 and 10000 ppm, copper hydroxide at 40, 60, 80 and 200 ppm, and magnesium chloride at 10000 ppm. Eggs were incubated in a flow through system at a density of 20 eggs·cm−2. After 61 days of incubation, the highest efficiencies were obtained with 200 ppm of copper hydroxide (77.6% of survivors to stage 2) with no significant differences from the control (74.3%). Lower concentrations of copper hydroxide resulted in high egg mortality whereas potassium sorbate and magnesium chloride were ineffective to avoid fungal growth and total egg mortality took place. From the obtained results, copper hydroxide baths at 200 ppm could be considered as a good alternative to formaldehyde.