Non-invasive quantification of lower airway antigen-specific antibodies using exhaled breath particles – an observational proof-of-concept study

Background Advancements in mucosal vaccine development highlight the urgent need for practical methods to assess immune responses in both upper and lower airways. Although nasal sampling is increasingly standardised, a non-invasive technique for directly assessing antigen-specific antibodies in the distal airways has been lacking. Methods We used Particles in Exhaled Air (PExA), a non-invasive method capturing aerosolised respiratory tract lining fluid (RTLF) from the distal airways, to quantify antigen-specific IgA and IgG. Antibodies were measured in paired serum, nasal secretions, and PExA samples from 28 healthcare workers during spring 2025, including 18 with RT-qPCR-confirmed SARS-CoV-2, influenza A (H1N1), or respiratory syncytial virus (RSV) infection. Findings Antigen-specific IgA and IgG measurements in PExA samples showed low variability (median CVs: spike-IgA 3.7%, H1-IgA 7.2%, RSV pre-F-IgA 5.3%). Strong correlations (Spearman's rho >0.85, p < 0.0001) were observed between PExA and serum IgG, which may be consistent with a systemic contribution to lower-airway IgG levels. In contrast, IgA correlations were more modest, which may reflect local IgA production in the lungs. IgG and IgA correlated between PExA and nasal secretions, while the IgA/IgG ratio was higher in nasal secretions (median 5.0 vs 0.6, p < 0.0001), consistent with expected isotype distribution in upper and lower airways. Interpretation This study demonstrates the feasibility of non-invasive measurement of antigen-specific antibodies in the lower airways. Our approach proposes a framework for assessing lower-airway immunity and supporting next-generation mucosal vaccine development. Funding This study was funded by the European Research Council, the Swedish Society for Medical Research, and the Knut and Alice Wallenberg Foundation.