Fig. 5
Toxin neutralization capacity of anti-rLTB.Etx40–62 antibodies: a in vitro neutralization ability of the anti-fusion protein antisera against the epsilon toxin was determined by incubating the rEtx (75 ng/ml) with different dilutions of anti-fusion protein antisera [undiluted (neat), 1:10, 1:25, 1:50, 1:100 and 1:500] prior to its addition to the MDCK cells. The cells were then incubated for 1 h at 37 °C and cell viability was determined by MTT assay. Cells treated with the rEtx pre-incubated with the undiluted pre-immune sera (PI, neat) were included as control. Percent survival of MDCK cells was plotted with respect to untreated control cells. Ordinary one-way ANOVA was performed to determine the statistically significant change (p value) in the survival with respect to rEtx-treated cells. **p ≤ 0.005; ***p ≤ 0.001. b BALB/c mice (n = 10 per group) were challenged with the 2 × LD50 dose of the rEtx (3 ng/20 g body weight in 5 µl PBS; intraperitoneal) pre-incubated with equal volume of undiluted anti-fusion protein antisera, generated with or without alum or preimmune serum (PI) at 37 °C for 1 h. The mice were monitored for survival for 5 days. Percentage survival was calculated according to the observed mortality