2.1 Antigen and animals
HEK293 expressing recombinant SARS-CoV-2 S1 of the original Wuhan strain (Assession# QHD43416.1, Val 16–Arg 685) was purchased from ACROBiosystems (Newark, DE). Female BALB/c mice (age 6-8 weeks) were purchased from Japan SLC (Shizuoka, Japan). All animals were maintained under specific-pathogen-free conditions. Mice were anesthetized by intraperitoneal injection of 62.6 mg ketamine and 12.4 mg xylazine per kg body weight before vaccination. All mice used in this study were treated according to the Guide for the Care and Use of Laboratory Animals (NIH Publication No. 85-23, 1996) and the study was approved by the Animal Care Committee of Tokushima University (#T2020-65).