The Omicron variant (B.1.1.529) of SARS-CoV-2, first detected in southern Africa, has rapidly spread globally due to its enhanced transmissibility and a large number of spike mutations. This study found that the Omicron variant is highly resistant to neutralization by serum from both convalescent patients and individuals vaccinated with widely used COVID-19 vaccines. Even serum from individuals who received booster doses of mRNA-based vaccines showed significantly reduced neutralizing activity against the Omicron variant. A panel of monoclonal antibodies targeting various epitope clusters on the spike protein revealed that 17 out of 19 antibodies tested lost or impaired their neutralizing activity against the Omicron variant. Additionally, four new spike mutations (S371L, N440K, G446S, and Q493R) were identified, which confer greater antibody resistance to the Omicron variant. These findings highlight the serious threat posed by the Omicron variant to existing COVID-19 vaccines and therapies, emphasizing the need for new interventions to anticipate the virus's evolutionary trajectory.The Omicron variant (B.1.1.529) of SARS-CoV-2, first detected in southern Africa, has rapidly spread globally due to its enhanced transmissibility and a large number of spike mutations. This study found that the Omicron variant is highly resistant to neutralization by serum from both convalescent patients and individuals vaccinated with widely used COVID-19 vaccines. Even serum from individuals who received booster doses of mRNA-based vaccines showed significantly reduced neutralizing activity against the Omicron variant. A panel of monoclonal antibodies targeting various epitope clusters on the spike protein revealed that 17 out of 19 antibodies tested lost or impaired their neutralizing activity against the Omicron variant. Additionally, four new spike mutations (S371L, N440K, G446S, and Q493R) were identified, which confer greater antibody resistance to the Omicron variant. These findings highlight the serious threat posed by the Omicron variant to existing COVID-19 vaccines and therapies, emphasizing the need for new interventions to anticipate the virus's evolutionary trajectory.