Fig. 6: Mpx-V3 immunization protects BALB/c mice from intranasal challenge of MPXV.
From: Next-gen novel nanocage-based multivalent vaccine candidate to tackle the rising menace of Mpox
35 female BALB/c mice of 6–8 weeks of age were randomly divided into five groups, having n = 7 mice in each group. a Following the previous immunization scheme and dose, mice were immunized in a prime and boost dose of 25 μg along with AddaVaxTM and Alhydrogel as shown in the schematic. b Neutralizing antibody titers (n = 6) of day 35 Mpx-V3 sera against VACV (WR), MPXV (N-27, from BeI resources), and MPXV (Clade IIb, from NIV Pune) were determined by the PRNT50 method. PRNT50 value from individual mice is plotted, and the error bar represents the SEM. c The immunized mice (n = 7) were intranasally challenged with 50 μL 1 × 106 PFU/mL of MPXV. Post-infection changes in body weight and clinical scores were recorded every day for 2 weeks. Animals were scored based on the clinical symptoms exhibited by them into 1–10. Scoring with 0 = no alteration in behavior, 1 = no restriction of movement; blink frequently; no body stiffening; no hind limb paralysis; 2 = dull; 3 = piloerection; 4 = shivering; 5 = hunched posture; 6 = restriction of movement; 7 = blink frequently; no body stiffening; no hind limb paralysis; 8 = restriction of movement; body stiffening; no hind limb paralysis; 9 = restriction of movement; eyes closed; 10 = body stiffening; hind limb paralysis, sometimes tremor. The data shown here is each group’s mean ± standard deviation. d The virus titer of MPXV was determined in the lung and trachea samples (n = 6) harvested at 6 days post-infection, respectively. Upon harvesting, organ samples were weighed, homogenized, sonicated, and centrifuged. Later, the aliquots of the supernatant from the samples were stored for titer calculation in Vero cells. Confluent Vero cells were infected with different dilutions of organ supernatant for 2 h, and later the cells were washed and layered with 2% CMC in 2% FBS containing DMEM for 48 h. Then, the media was removed, cells were fixed with 4% paraformaldehyde, and stained with 0.5% crystal violet. Plaques were counted from the plates to calculate virus titer in PFU/g. The values are plotted as mean ± SEM, and statistical analysis between the two groups was performed by unpaired, two-tailed Student’s t-test. GraphPad Prism 9 was used for calculating statistical analysis. One-way ANOVA was used for comparing multiple groups, and Dunnett’s multiple comparison tests were applied to calculate the p values.
