Fig. 2
Assessment and comparison of IgG and IgG subclass antibodies to rPfCelTOS in the sera of vaccinated mice by ELISA (a) and Th1/Th2 responses ratios (b). Groups of 6- to 8-week-old female BALB⁄c mice were immunized subcutaneously with rPfCelTOS alone or formulated in different adjuvants: CpG, Poly I:C, CpG + Poly I:C, and CFA/IFA (as the reference adjuvant). Anti-rPfCelTOS IgG and its subclasses were assessed in individual immunized mice on day 38 of the first immunization (a). IgG2a was the predominant isotype in all vaccine groups (1–5). The highest levels of IgG2a and IgG2b subclasses were detected in the group received rPfCelTOS in combination with CpG + Poly I:C (P < 0.0001, one-way ANOVA). The bars and error bars show the mean OD490nm and SD of 16 individual mice in each group, respectively. The ELISA cut-offs were calculated as the mean OD490nm of NMS (as the negative controls, n = 20) plus 3SD. The cut-off values for total IgG, IgG1, IgG2a, IgG2b, and IgG3 antibodies were 0.101, 0.064, 0.072, 0.093, and 0.091, respectively. The Th1/Th2 ratio (IgG2a/IgG1, IgG2b/IgG1 and IgG2a + IgG2b/IgG1 + IgG3) was performed in the sera of immunized mice on day 38 after the first immunization (b) and among all vaccinated groups; the highest and a significant difference in all ratios was observed in group 4, rPfCelTOS/CpG + Poly I:C (P < 0.0001, one-way ANOVA). Data was analysed using one-way ANOVA followed by Tukey’s HSD post hoc test. *P < 0.05, **P < 0.001, ***P < 0.0001. Ag rPfCelTOS antigen, TIgG total IgG, NMS normal mouse sera