*2.6. Animal Challenge Studies*

For the Zika challenge studies, IFNAR−/<sup>−</sup> mice (*n* = 12–14/group) were immunized once with 0.5 μg of Zika prME vaccine or 0.5 μg of Zika prME and 11 μg of opt-36γt. The mice were challenged with 1 <sup>×</sup> 105 PFU ZIKV-PR209 virus via intraperitoneal (IP) injection on day 15. Post challenge, the animals were weighed daily. In addition, they were observed for clinical signs of disease twice daily (decreased mobility; hunched posture; hind-limb knuckle walking (partial paralysis); and/or paralysis of one hind limb or both hind limbs). The criteria for euthanasia on welfare grounds consisted of 20% weight loss or prolonged paralysis in one or both hind limbs.
