For S, it was decided to use the stabilized trimer (R683A, R685A, K986P, V987P), since participating teams had also modeled trimer-specific or cryptic binding sites, other than the (classical) RBD domain. As a positive control, the natural choice was the Ace2 (human receptor) protein. Surprisingly, recombinantly expressed Ace2 did not show binding to S (stabilized trimer), we suspect due to improper folding of the construct. Fortunately, His-tagged Ace2 did provide good binding curves with a K D of 4.25 ± 1.52 nM (over 6 runs performed during the 3 days of assay measurements, see Supporting Information Section 4.3.2). This is stronger binding than previous measurements performed by Surface Plasmon Resonance 76 that showed 94.6 ± 6.5 nM for (monomeric) SARS-CoV-2-S1, but can be explained by multivalency of the trimer as shown by Kruse et al. . All 152 compounds were first analyzed using 8-point dilution series between 50 nM and 100 µM concentrations, revealing 7 compounds to be potential binders. The latter 7 were measured in triplicate 12-point dilutions from 0.2 nM to 200 µM, and 3 compounds were identified as high micromolar binders: S-1, S-2 and S-3 (see Figure and table 3 below).