BACKGROUND: Seroreactivity against human endemic coronaviruses has been linked to disease MESHD severity after SARS-CoV-2 infection MESHD. Assays that are capable of concomitantly detecting antibodies SERO against endemic coronaviridae such as OC43, 229E, NL63, and SARS-CoV-2 may help to elucidate this question. We set up a platform for serum SERO-screening and developed a bead-based Western blot system, namely DigiWest, capable of running hundreds of assays using microgram amounts of protein prepared directly from different viruses. METHODS: The parallelized and miniaturised DigiWest assay was adapted for detecting antibodies SERO using whole protein extract prepared from isolated SARS-CoV-2 virus particles. After characterisation and optimization of the newly established test, whole virus lysates of OC43, 229E, and NL63 were integrated into the system. RESULTS: The DigiWest-based immunoassay SERO system for detection of SARS-CoV-2 specific antibodies SERO shows a sensitivity SERO of 87.2 % and diagnostic specificity of 100 %. Concordance analysis with the SARS-CoV-2 immunoassays SERO available by Roche, Siemens, and Euroimmun indicates a comparable assay performance SERO (Cohen's Kappa ranging from 0.8799-0.9429). In the multiplexed assay, antibodies SERO against the endemic coronaviruses OC43, 229E, and NL63 were detected, displaying a high incidence of seroreactivity against these coronaviruses. CONCLUSION: The DigiWest-based immunoassay SERO, which uses authentic antigens from isolated virus particles, is capable of detecting individual serum SERO responses against SARS-CoV-2 with high specificity and sensitivity SERO in one multiplexed assay. It shows high concordance with other commercially available serologic assays. The DigiWest approach enables a concomitant detection of antibodies SERO against different endemic coronaviruses and will help to elucidate the role of these possibly cross-reactive antibodies SERO.