The study investigates how the SARS-CoV-2 Omicron variant, which contains 15 mutations in the receptor-binding domain (RBD), escapes neutralizing antibodies. Using high-throughput yeast display screening, the researchers classified 247 human anti-RBD neutralizing antibodies into six epitope groups (A–F) and found that various single mutations of Omicron can impair neutralizing antibodies from different groups. Specifically, antibodies in groups A–D, which overlap with the ACE2-binding motif, are largely escaped by K417N, G446S, E484A, and Q493R. Antibodies in groups E and F, which often exhibit broad sarbecovirus neutralizing activity, are less affected but still show some escape by G339D, N440K, and S371L. The study also found that neutralizing antibodies that tolerate single mutations can be further impaired by multiple synergistic mutations on their epitopes. Overall, over 85% of the tested neutralizing antibodies were escaped by Omicron. Neutralizing antibody-based drugs, such as LY-CoV016, LY-CoV555, REGN10933, REGN10987, AZD1061, AZD8895, and BRRI-196, were significantly reduced in potency by Omicron, while VIR-7831 and DXP-604 still functioned at a reduced efficacy. The findings suggest that Omicron could cause substantial humoral immune evasion and that neutralizing antibodies targeting the conserved region of sarbecovirus will remain most effective.The study investigates how the SARS-CoV-2 Omicron variant, which contains 15 mutations in the receptor-binding domain (RBD), escapes neutralizing antibodies. Using high-throughput yeast display screening, the researchers classified 247 human anti-RBD neutralizing antibodies into six epitope groups (A–F) and found that various single mutations of Omicron can impair neutralizing antibodies from different groups. Specifically, antibodies in groups A–D, which overlap with the ACE2-binding motif, are largely escaped by K417N, G446S, E484A, and Q493R. Antibodies in groups E and F, which often exhibit broad sarbecovirus neutralizing activity, are less affected but still show some escape by G339D, N440K, and S371L. The study also found that neutralizing antibodies that tolerate single mutations can be further impaired by multiple synergistic mutations on their epitopes. Overall, over 85% of the tested neutralizing antibodies were escaped by Omicron. Neutralizing antibody-based drugs, such as LY-CoV016, LY-CoV555, REGN10933, REGN10987, AZD1061, AZD8895, and BRRI-196, were significantly reduced in potency by Omicron, while VIR-7831 and DXP-604 still functioned at a reduced efficacy. The findings suggest that Omicron could cause substantial humoral immune evasion and that neutralizing antibodies targeting the conserved region of sarbecovirus will remain most effective.