Tehran University of Medical Sciences Journal of Arthropod-Borne Diseases 2322-1984 16 1 2022 11 14 1 12 Narges Safari-Foroushani Islamic Azad University, Science and Research Branch, Tehran, Iran Mohammad Hossein Modarressi Department of Medical Genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran Kamran Pooshang Bagheri Venom and Bio therapeutics Molecules Lab, Medical Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran Mahdi Behdani Venom and Bio therapeutics Molecules Lab, Medical Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran Delavar Shahbazzadeh Venom and Bio therapeutics Molecules Lab, Medical Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran 2021 06 15 2022 11 02 Background: Hemiscorpius lepturus is one of the most dangerous scorpions in Iran and the world. Numerous studies have been conducted on phospholipases, especially phospholipase D, in this scorpion’s venom, and the results have shown this protein to be the main cause of death. Therefore, one of the most effective ways of preventing fatalities is to produce a toxoid vaccine from the deadly toxin of the venom. The present study was conducted to assess the non-toxicity of this toxoid and the safety of the vaccine candidate in BALB/c mice. Methods: The production of interferon-gamma and interleukin-4 cytokines in the spleen cells of the mice was measured using ELISpot assay 28 days following immunization with rPLD toxoid. Results: The unpaired t-test results showed a significant increase in the concentration of IFN-γ cytokine in the vaccinat­ed mice (P= 0.001), indicating that the immune system is directed toward the Th1 pattern, while no significant differ­ence was observed in the levels of IL-4 (P= 0.16) despite an increase in this cytokine. The in-vivo tests showed that the mice immunized with interval doses of 80µg of toxoid were completely protected against 10 × the LD100 of the venom. Moreover, the toxoid had no dermonecrotic effects and caused no necrotic and inflammatory complications in the rabbit skin. Conclusion: As a vaccine, the toxoid has the potential to increase the Th1 cytokine response and, subsequently, in­crease acquired cellular immunity. Thus, this toxoid appears to be able to provide an effective vaccine against the ven­om of Hemiscorpius lepturus. https://jad.tums.ac.ir/index.php/jad/article/view/1455 https://jad.tums.ac.ir/index.php/jad/article/download/1455/581