Allergy to Myrmecia pilosula (Jack Jumper Ant) venom is common in Australia, affecting ∼2.7% of some communities. Venom immunotherapy is a highly effective treatment, but for the venom to be widely distributed for clinical use, the stability and shelf-life of formulated Jack Jumper Ant venom must be demonstrated. HPLC-UV, ELISA Inhibition, SDS-PAGE and SDS-PAGE Immunoblot were used to assess venom stability under conditions of varying temperature, pH and in the presence of various stabilising agents. Optimal stability occurred between pH 8 and 10, however the presence of benzyl alcohol within this pH range resulted in a cloudy appearance within 3 days, so a pH of 6 was used. Increasing polysorbate 80 concentrations accelerated the degradation of allergenic peptides in 100 μg/mL venom, but improved stability at concentrations of 1 μg/mL or less. Sucrose reduced degradation of allergens Myr p 1 and Myr p 3, whilst glycerol was destabilising. In the presence of 22% sucrose, 1.1. mg/mL Jack Jumper Ant venom was stable at -18 °C and 4 °C for 12 months; following dilution to 100 μg/mL with 0.9% sodium chloride, 10. mM phosphate (pH 6), 0.05% polysorbate 80 and 0.9% benzyl alcohol (giving 2% sucrose), venom was stable for 7 days when stored at 4 °C. Concentrated Jack Jumper Ant venom can be stored in 22% sucrose for 12 months, and after dilution to 100 μg/mL for clinical use, it should be discarded after 7 days.