Total specific IgG isotype levels in immunised mice with different vaccine formulations. The sera of immunised mice were collected 0, 15, 30, 45, and 60 days post-immunisation and tested by ELISA for the presence of specific IgG1 (A), IgG2a (B) and IgG3 (C). The results are presented as the means and standard deviation (bars) for each experiment with ten animals per group. Significant differences between IgG production in the different groups was calculated employing one-way ANOVA followed by the Tukey post-test. D,d: groups significantly different on day 30 (p < 0.05); E,e: groups significantly different on day 45 (p < 0.05); F,f: groups significantly different on day 60 (p < 0.05). Arrows correspond to the two immunisations (days 0 and 15), and stars indicate the challenge with the virulent strain (Day 30). All bleedings were performed before immunisations and challenge.