The ongoing evolution of SARS-CoV-2, particularly through the emergence of new variants, continues to challenge our understanding of immune protection. While antibody levels correlate with protection against earlier variants like Alpha and Delta, their relationship with Omicron sub-variants remains unclear. To investigate the role of antibody levels and neutralizing activity in preventing breakthrough infections, we analyzed longitudinal SARS-CoV-2 humoral responses and neutralizing activity against the ancestral virus and major emerging variants in a well-characterized cohort of healthcare workers in Spain (N = 405). We find that antibody levels and neutralization titers are key indicators of protection against SARS-CoV-2, including the more evasive BQ.1 and XBB Omicron variants. Higher IgG and IgA levels are associated with protection over three 6-month follow-up periods sequentially dominated by BA.1, BA.2, BA.5, BQ.1, and XBB Omicron sub-variants, although the strength of the association between antibody levels and protection declines over time. Our findings demonstrate that binding antibody levels and neutralizing responses are a valid correlate of protection against more evasive BQ.1 and XBB Omicron variants, although the strength of this association diminishes over time. Additionally, our results underscore the importance of continuous monitoring and updating vaccination strategies to maintain effective protection against emerging SARS-CoV-2 variants.