Production of a Bacillus anthracis Secretome with Suitable Characteristics as Antigen in a Complement Fixation Test

In this study, we cultured the Bacillus anthracis vaccine strain Sterne 34F2 in a medium containing EDTA, and we assessed the best conditions to inhibit the activity of zinc-dependent metalloproteases to obtain a secretome containing a high concentration of non-degraded PA (PA(83)), as evaluated by the SDS-PAGE analysis. Then, we used this secretome as the antigen in a Complement Fixation Test (CFT) to monitor the production of antibodies against PA(83) in the sera of rabbits vaccinated with Sterne 34F2 and then infected with a B. anthracis virulent strain to evaluate the potency of the vaccine. The PAS-based CFT results were compared with those obtained by using a commercial ELISA kit. The two serological tests gave similar results in terms of specificity and sensitivity, as the kinetics of the antibodies production was very similar. The Sterne 34F2 vaccine induced an antibody response to PA(83), whose titer was not inferior to 1:8 in PAS-based CFT and 42 kU/mL in PA(83)-based ELISA, respectively, in all vaccinated rabbits. Our opinion is that the PAS-based CFT can be successfully employed in humans and in animals for epidemiological retrospective studies or post-vaccination monitoring. We also suggest the use of our method to test the efficacy of veterinary anthrax vaccines.