Per-and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants that are widely detected in human blood samples. Recent epidemiological studies suggest a negative correlation between PFAS blood concentrations and SARS-CoV-2 antibody responses following COVID-19 vaccination. Despite these observations, the direct impact of PFAS mixtures on the interaction between circulating T follicular helper (cTfh) and B cells, which is pivotal for the production of neutralizing antibodies in response to SARS-CoV-2 vaccination, remains unexplored. Using human peripheral blood mononuclear cells (PBMCs) isolated from six healthy female donors who had previously been vaccinated and subsequently boosted or recovered from COVID-19, we examined in vitro the effect of a PFAS mixture on cTfh and B-cell immune responses to SARS-CoV-2 spike peptide. The PBMCs were exposed to a PFAS mixture comprising perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonate (PFHxS), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluoroundecanoic acid (PFUnDA) at human-relevant blood concentrations (1X), and higher concentrations (10X, 100X, 1000X), followed by stimulation with SARS-CoV-2 spike peptides for five days. We showed that the PFAS mixtures did not induce cytotoxicity but caused immunomodulation. Exposure to the highest PFAS concentration tested (1000X human blood concentrations) led to a selective reduction in cTfh17 cells and upregulation of ICOS and IL-21 in cTfh1/17 cells. B cells exhibited unusual activation, with naïve B cells showing elevated CD80/CD86 expression and switched memory B cells upregulating CD69. Furthermore, despite unperturbed levels of IL-21 and CXCL13, important mediators involved in antibody production, we observed a significant reduction in plasmablasts and SARS-CoV-2 spike-specific IgG secretion at the highest PFAS concentrations. In conclusion, this study provides in vitro evidence that PFAS mixtures can modulate selected components of the cTfh-B-cell axis and may contribute to impaired vaccine-induced antibody responses under high-exposure conditions. These findings add an important piece of information as to why high PFAS-exposed individuals present reduced COVID-19 vaccine antibody titers.