Simultaneous In Situ Visualization and Quantitation of Dual Antigens Adsorbed on Adjuvants Using High Content Analysis

Aim: Traditional antigenicity assay requires antigen recovery from the particulate adjuvants prior to analysis. An in situ method was developed for interrogating vaccine antigens with monoclonal antibodies while being adsorbed on adjuvants. Materials & methods: The fluorescence imaging-based high content analysis was used to visualize the antigen distribution on adjuvant agglomerates and to analyze the antigenicity for adsorbed antigens. Results: Simultaneous visualization and quantitation were achieved for dual antigens in a bivalent human papillomavirus vaccine with uniquely labeled antibodies. Good agreement was observed between the in situ multiplexed assays with well-established sandwich enzyme-linked immunosorbent assays. Conclusion: The streamlined procedures and the amenability for multiplexing make the in situ antigenicity analysis a favorable choice for in vitro functional assessment of bionanoparticles as vaccine antigens.