Purification of Anti-SARS-CoV-2 Antibodies

Anti-S-IgGs and anti-RBD-IgGs were obtained using S-Sepharose and RBD-Sepharose on an Akta Start chromatograph (GE Life Sciences, New York, NY, USA) and as described in [46 (link)]. The sorbents with immobilized S-protein and RBD were prepared according to the standard protocol using CNBr Sepharose (GE Life Sciences, New York, NY, USA) and our previously published works [46 (link),63 (link)]. An equimolar mixture of 10 IgG preparations was applied to 3 mL of RBD-Sepharose pre-equilibrated with 50 mM Tris-HCl, pH 7.5, containing 0.15 M NaCl (TBS). The fraction eluted with the acidic buffer was neutralized by adding 1/10 v/v 1.0 M Tris-HCl, pH 8.8, and then dialyzed against 20 mM Tris-HCl, pH 7.5. The IgG fraction not bound to RBD-Sepharose was applied to a 10 mL S-Sepharose column pre-equilibrated with TBS. The IgGs were eluted with a two-step gradient: 50 mM Tris-HCl, pH 7.5, containing 1.0 M NaCl and 0.1 M Gly-HCl, pH 2.6. The resulting fraction was also dialyzed against 20 mM Tris-HCl, pH 7.5.