Trivalent P-type S60-VP8* Nanoparticle Vaccine

Three P-types of S₆₀-VP8* nanoparticles (P[8], P[6], and P[4]) were produced and purified as previously described [31 (link),33 (link)] and mixed as the trivalent vaccine in the liquid formulation in phosphate buffer saline (PBS). The individual nanoparticles and the trivalent vaccine were assessed by (1) SDS-PAGE for protein quality; (2) gel filtration chromatography and EM inspection for nanoparticle formation and structural integrity [31 (link),33 (link)]. Bacterial endotoxin was removed by using ToxinEraser™ Endotoxin Removal Resin (GenScript, Cat. No L00402. Piscataway, NJ, USA). The vaccines were formulated by mixing each S₆₀-VP8* P[8], P[6], and P[4] or S₆₀ alone with 600 µg/dose of aluminum hydroxide adjuvant (Alhydrogel, InvivoGen, San Diego, CA, USA). The vaccine dose was originally designed to contain 200 µg of each P-type. However, due to the need for filtration to eliminate contaminating bacteria, the actual amount of antigen in the vaccine was reduced. The actual amount of antigens in the vaccine measured with Nanodrop was 94 µg of P[4], 52 µg of P[6], and 143 µg of P[8]. Sterility of antigens was confirmed using blood agar and thioglycolate broth prior to inoculation of Gn pigs.