BLI-based competitive assay
The ability of the sera of the COVID convalescent patient to compete with ACE2 for binding to RBDWT and RBDTRIP was tested in a sandwich format assay on the Octet RED96e (Fortebio). Anti-penta-His (HIS1K) biosensors were loaded for 10 min with RBDWT and RBDTRIP at a concentration of 7.5 µg/ml in BLI assay buffer followed by addition of samples (diluted 1:20 in BLI assay buffer) from convalescent human sera. To assess whether the sera can inhibit the binding of ACE2 to RBDWT and RBDTRIP, ACE2-mFc (50 nM) was added to biosensor. For control two additional sensors with BLI buffer were used, one for baseline and one without serum sample to determine binding of ACE2-mFc alone. The results are expressed of single individual. The response data were normalized using ForteBio data analysis software version1.2.0.1.55.