Characterization and evaluation of the enzymatic activity of tetanus toxin submitted to cobalt-60 gamma radiation

Abstract

Background: Tetanus toxin blocks the release of the inhibitory neurotransmitters in the central nervous system and causes tetanus and its main form of prevention is through vaccination. The vaccine is produced by inactivation of tetanus toxin with formaldehyde, which may cause side effects. An alternative way is the use of ionizing radiation for inactivation of the toxin and also to improve the potential immunogenic response and to reduce the post-vaccination side effects. Therefore, the aim of this study was to characterize the tetanus toxin structure after different doses of ionizing radiation of 60 Co. Methods: Irradiated and native tetanus toxin was characterized by SDS PAGE in reducing and non-reducing conditions and MALD-TOF. Enzymatic activity was measured by FRET substrate. Also, antigenic properties were assessed by ELISA and Western Blot data. Results: Characterization analysis revealed gradual modification on the tetanus toxin structure according to doses increase. Also, fragmentation and possible aggregations of the protein fragments were observed in higher doses. In the analysis of peptide preservation by enzymatic digestion and mass spectrometry, there was a slight modification in the identification up to the dose of 4 kGy. At subsequent doses, peptide identification was minimal. The analysis of the enzymatic activity by fluorescence showed 35 % attenuation in the activity even at higher doses. In the antigenic evaluation, anti-tetanus toxin

Advances in Irradiated Livestock Vaccine Research and Production Addressing the Unmet Needs for Farmers and Veterinary Services in FAO/IAEA Member States

“…The best way to currently prevent tetanus is through vaccination using formaldehyde inactivated tetanus toxin ( 52 ). In order to overcome the disadvantages of exposing vaccine recipients to long term effects of formaldehyde and associated salts, irradiation was used to inactivate tetanus toxin ( 53 ). The toxin was inactivated at 5 kGy but retained immunogenicity at 8 KGy which was the upper limit of irradiation used in the experiment.…”

“…In addition, pure irradiated toxin retained more than 50% of its enzymatic activity. Future studies will optimize the production process, detoxification and explore its feasibility as an adjuvant ( 53 ). Other toxin producing pathogens tested in irradiation studies include Mycobacterium spp., Anthrax, Cholera, Coli and paratyphoid B where irradiation does not necessarily inactivate the toxins in contaminated meat ( 28 , 54 – 56 ).…”