The safety and immunogenicity of a protein-based tetravalent vaccine SCTV01E that contains spike protein ectodomain (S-ECD) of Alpha, Beta, Delta and Omicron BA.1 are assessed and compared with bivalent protein vaccine SCTV01C (Alpha and Beta variants) and monovalent mRNA vaccine (NCT05323461). The primary endpoints are the geometric mean titers (GMT) of live virus neutralizing antibodies (nAb) to Delta (B.1.617.2) and Omicron BA.1 at day 28 post-injection. The secondary endpoints include the safety, day 180 GMTs against Delta and Omicron BA.1, day 28 GMTs to BA.5, and seroresponse rates of neutralizing antibodies and T cell responses at day 28 post-injection. 450 participants, comprising of 449 males and 1 female, with a median age (range) of 27 (18–62) years, are assigned to receive one booster dose of BNT162b2, 20 µg SCTV01C or 30 µg SCTV01E and completed 4-week follow-up. All SCTV01E related adverse events (AEs) are mild or moderate and no Grade ≥3 AE, serious AE or new safety concerns are identified. Day 28 GMT of live virus neutralizing antibodies and seroresponse against Omicron BA.1 and BA.5 with SCTV01E are significantly higher than those with SCTV01C and BNT162b2. These data indicate an overall neutralization superiority with tetravalent booster immunization in men.