As01b

8-week-old female C57BL6/J mice (Jackson Lab, US) were vaccinated subcutaneously in the back of neck at day 0 (Prime) and day 14 (Boost) using a vaccine including 5 μg recombinant gE protein (GSK, UK) and indicated adjuvant (AS01b: 50 μL/mouse, AS-QM40be: 50 μL/mouse, AS-QM40cc: 50 μL/mouse, AS-M40: 50 μL/mouse, Addavax: 50 μL/mouse, Alhydrogel: 50 μL/mouse). AS01b and gE were from GSK Shingrix vaccine. AS-QM40be, AS-QM40cc, AS-M40 were in-house products at Agenus/SaponiQx as describe above. Addavax and Alhydrogel were from Invivogen. Plasma was collected at day 0, 14, 21, 28 for anti-gE IgG2c/IgG1/IgGs ELISA detection. Splenocytes were harvested at day 28 for Elispot and Luminex cytokines analysis.

Six-week-old female specific pathogen-free (SPF) C57BL/6N mice (15–18 g) were purchased from Vital River Laboratory Animal Technology Ltd. (Beijing, China), randomly divided into groups of 6 mice each (n = 6), maintained under SPF conditions and housed with free access to food and water at the Central Animal Services of the Institute of Medical Biology, Chinese Academy of Medical Sciences (IMB, CAMS). Polyinosinic–polycytidylic acid (Poly I:C, from InvivoGen, Inc., San Diego, CA, USA) and alum (Thermo Fisher, Eugene, OR, USA) mixtures or AS01B (from GSK, MD, USA)-adjuvanted extracellular domain of gE were used as controls, and PBS was used as a blank control. The mice were immunized intramuscularly in the thigh muscle twice with 50 μL of immunogen at 3-week intervals. After anesthetization by intraperitoneal injection of tribromoethanol, blood samples (via cardiac puncture) and spleens were collected 2 weeks after the final immunization for further analysis.