Analysis of the secondary structure of RBD by CD spectroscopy and verification of antigenic intactness
(A) Representative CD spectral data as analyzed by online server CAPITO. (B) SDS-PAGE analysis of the purified RBD. RBD was purified using anti-FLAG resin from RAW cells infected with Salmonella carrying pJHL204-FLAG-RBD. (C) Western blot and (D) ELISA results demonstrate reactivity of purified RBD against a known spike neutralizing mAb from the mouse (clone 57, RBD-mFc immunogen). Lane M, protein molecular weight marker; lane S1 and S2, respectively, denote purified protein samples 1 and 2. For ELISA, serial dilutions of RBD MAb ranging from 102 to 105 were reacted with purified RBD for 1 h at 37°C. Data in (D) represent mean ± SEM of OD492 values derived from two protein sample preparations. pJHL204, SFV replicon-based vector.