Coronavirus is an enveloped RNA virus that causes mild to severe
respiratory diseases in humans, including HKU1-CoV, 229E-CoV, NL63-CoV, OC43-CoV, SARS-CoV, MERS-CoV, and SARS-CoV-2. Due to the outbreak of SARS-CoV-2, it is important to identify the patients and investigate their immune responses.
Protein microarray is one of the best platforms to profile the
antibodies in the blood because of its fast, multiplexed, and sensitive nature. To fully understand the immune responses and biological specificities, this study developed a human coronavirus (HCoV)
protein microarray and included all seven human coronaviruses and three influenza viruses. Each
protein was printed in triplicate and formed 14 identical blocks per array. The HCoV
protein microarray showed high reproducibility and sensitivity to the
monoclonal antibodies against spike and
nucleocapsid protein with detection limits of 10-200 pg. The HCoV
proteins that were immobilized on the array were properly folded and functional by showing interactions with a known human receptor, e.g., ACE2. By profiling the serum
IgG and
IgA from 32
COVID-19 patients and 36 healthy patients, the HCoV
protein microarray demonstrated 97% sensitivity and 97% specificity with two
biomarkers. The results also showed the cross-reactivity of
IgG and
IgA in
COVID-19 patients to spike
proteins from various coronaviruses, including that from SARS-CoV, HKU1-CoV, and OC43-CoV. Finally, an innate immune
protein named
surfactant protein D showed broad affinities to spike
proteins in all human coronaviruses. Overall, the HCoV
protein microarray is multiplexed, sensitive, and specific, which is useful in diagnosis, immune assessment, biological development, and drug screening.