Science 367, 1260C1263 (2020). A SARS-CoVCderived RBD from palm civets elicited more potent cross-neutralizing responses in immunized animals than the RBD from a human SARS-CoV strain, informing strategies for development of universal vaccines against emerging coronaviruses. INTRODUCTION The global outbreak of the coronavirus disease 2019 (COVID-19) was caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is a new coronavirus (CoV) genetically close to SARS-CoV that emerged in 2002 ( 0.01 and *** 0.001. Mouse and rabbit antisera developed against SARS-CoV RBD cross-react and neutralize SARS-CoV-2 As the S protein RBD dominates the nAb response to SARS-CoV, we sought to characterize the RBD-mediated cross-reactivity and neutralization on SARS-CoV-2. To this end, we first generated mouse anti-RBD sera by immunization with two RBD-Fc fusion proteins: one encoding the RBD sequence of a palm civet SARS-CoV strain SZ16 (SZ16-RBD) and the second one with the RBD sequence of a human SARS-CoV strain GD03 (GD03-RBD). Both the fusion proteins were expressed in 293T cells and purified to apparent homogenicity (fig. S1). As shown in Fig. 4A, all eight mice developed strong antibody responses against the SARS-CoV S1 and RBD, and in comparison, four mice (M-1 to M-4) immunized with SZ16-RBD exhibited higher titers of antibody responses than the mice (M-5 to M-8) immunized with GD03-RBD. Each of the anti-RBD sera cross-reacted well with the S protein of SARS-CoV-2, suggesting that SARS-CoV and SARS-CoV-2 Amyloid b-peptide (42-1) (human) do share antigenically conserved epitopes in the RBD sites. Noticeably, while the SZ16-RBD immune sera also reacted with the SARS-CoV-2 S1 and RBD antigens, the cross-reactivity of the GD03-RBD immune sera was low. However, while the mouse anti-RBD sera at 1:50 dilutions were measured with increased coating antigens in ELISA, they reacted with the SARS-CoV-2 S1 and RBD efficiently, which verified the cross-reactivity (Fig. 4B). Similarly, the neutralizing activity of mouse antisera was determined by pseudovirus-based single-cycle contamination assay. As shown in Fig. 4 (C and D), both the SZ16-RBDC and GD03-RBDCspecific antisera Amyloid b-peptide (42-1) (human) displayed very potent activities to neutralize SARS-CoV; they also cross-neutralized SARS-CoV-2 with relatively lower efficiencies. As judged by the neutralizing activity at the highest serum dilution, the SZ16-RBD antisera were more potent than the GD03-RBD antisera in neutralizing SARS-CoV; however, the two antisera had no significant difference in neutralizing SARS-CoV-2 (Fig. 4, E Amyloid b-peptide (42-1) (human) and F). Open in a separate window Fig. 4 Cross-reactive and neutralizing activities of antisera from mice immunized with the RBD proteins of SARS-CoV.(A) Binding activity of mouse antisera at a 1:100 dilution to SARS-CoV (S1 and RBD) and SARS-CoV-2 (S, S1, and RBD) antigens was determined by ELISA. A healthy mouse serum was tested as control. (B) The cross-reactivity of mouse antisera with the SARS-CoV-2 S1 and RBD proteins. The antisera Amyloid b-peptide (42-1) (human) were diluted at 1:50, and the S1 and RBD antigens were coated at 100 ng per ELISA plate well. (C and D) Neutralizing activities of mouse antisera at indicated dilutions against SARS-CoV, SARS-CoV-2, and VSV-G pseudoviruses were determined by a single-cycle contamination assay. The experiments were performed in triplicate and repeated three times, and data are shown as means with SDs. (E and F) Comparison of neutralizing activities of the mouse antiCSZ16-RBD and antiCGD03-RBD sera. Statistical significance was tested by two-way ANOVA with Dunnett posttest. ns, not significant. * 0.05, ** 0.01, and *** 0.001. We further developed rabbit antisera by immunizations, in which two rabbits were immunized with SZ16-RBD (R-1 and R-2) or with GD03-RBD (R-3 and R-4). Each RBD protein elicited antibodies highly reactive with both the SARS-CoV and SARS-CoV-2 antigens (Fig. 5A), which were different from MGC4268 their immunizations in mice. As expected, all of the rabbit antisera potently neutralized SARS-CoV and SARS-CoV-2 in a Amyloid b-peptide (42-1) (human) similar profile with that of the mouse anti-S and anti-RBD sera (Fig. 5, B and C). Obviously, the neutralizing activity of rabbit antiCSZ16-RBD sera against both the viruses was higher than that of the rabbit antiCGD03-RBD sera (Fig. 5, D and E). Together, the results verified that this SARS-CoV S protein and its RBD immunogens can induce cross-neutralizing antibodies toward SARS-CoV-2 by vaccination. Open in a separate window Fig. 5 Cross-reactive and neutralizing activities of antisera from rabbits immunized with the RBD proteins of SARS-CoV.(A) Binding activity of rabbit antisera at a 1:100 dilution to SARS-CoV (S1 and RBD).