codon optimized plasmid cmv r sars cov 2 encoding d614g spike variant  (ATCC)

Journal: Vaccines

Article Title: Newcastle Disease Virus-Like Particles Displaying Prefusion-Stabilized SARS-CoV-2 Spikes Elicit Potent Neutralizing Responses

doi: 10.3390/vaccines9020073

Figure Lengend Snippet: Single immunization of S2P-NDVLP elicited a high immune response against SARS-CoV-2 virus in mice at Week 2. ( A ) Scheme of the immunization regimen and groups. The control immunogen was a trimeric spike S2P glycoprotein. The amount of S2P in the S2P-NDVLP immunogen was quantified by Western blotting with S2P-immunized mouse sera (see ). ( B ) S2P-NDVLP elicited substantial IgG titers against the SARS-CoV-2 spike at Week 2. The endpoint IgG titers were measured, and EC 50 values were calculated as the readout for ELISAs with Prism. The triangle symbol indicates the ELISA titer at assay maximum. ( C ) S2P-NDVLP elicited substantial SARS-CoV-2 pseudovirus neutralization titers at Week 2. Serum-neutralizing antibodies against a D614G pseudovirus were measured, and ID 50 values were calculated as the readout of the neutralization assay with Prism. The geometric mean titer (GMT) of each group is marked in red numbers on the horizontal axis. ( D ) Correlation between neutralization titers and ELISA S2P-specific IgG titers existed in the S2P-NDVLP-immunized groups but not in the S2P-immunized groups. ( E ) RBD-specific serum antibody titers were measured as BLI responses and calculated with Prism. The titers among the S2P-NDVLP-immunized groups were dose dependent at Week 2. ( F ) Correlation between neutralization titers and BLI RBD-specific antibody titers existed in the S2P-NDVLP-immunized groups but not in the S2P-immunized groups. ( G ) Quality of immune responses elicited by immunization was calculated as the ratio of neutralization titer to total ELISA IgG titer, plotted as mean and SD. The horizontal dotted line in ( C – F ) represents the low detection limit for the neutralization titer. In ( B – E ), the Kruskal–Wallis test was used to test differences between the groups, and P-values were designated as *: p < 0.05; **: p < 0.01; ***: p < 0.001; ****: p < 0.0001. Geometric means were indicated with a red bar. Spearman correlation was used in ( D , F ).

Article Snippet: A codon-optimized plasmid CMV/R-SARS-CoV-2-encoding D614G spike variant was constructed and cotransfected with a lentivirus backbone, luciferase reporter, and human transmembrane protease serine 2 (TMPRSS2) in HEK293T/17 cells (ATCC #CRL-11268).

Techniques: Virus, Control, Western Blot, Enzyme-linked Immunosorbent Assay, Neutralization

Journal: Vaccines

Article Title: Newcastle Disease Virus-Like Particles Displaying Prefusion-Stabilized SARS-CoV-2 Spikes Elicit Potent Neutralizing Responses

doi: 10.3390/vaccines9020073

Figure Lengend Snippet: S2P-NDVLP elicited a robust anti-SARS-CoV-2-neutralizing response with a relatively low S2P-equivalent dose in mice at Week 5. ( A ) Boost immunization elicited higher ELISA titers against the SARS-CoV-2 spike at Week 5. The titers among the S2P-NDVLP-immunized groups were not significantly different. IgG titers in the S2P-immunized groups were significantly higher than the S2P-NDVLP-immunized groups. Triangle symbols indicate the ELISA titers at assay maximum. ( B ) The S2P-NDVLP groups had robust neutralization activities at Week 5 but were not significantly different from each other. Serum neutralization titers (ID 50 ) were measured against a SARS-CoV-2 D614G variant pseudovirus. ( C ) Correlation between neutralization titers and ELISA S2P-specific IgG titers did not exist in the S2P-NDVLP-immunized groups, but a weak correlation existed in the S2P-immunized groups. ( D ) RBD-specific serum antibody titers were measured as BLI responses and calculated with Prism. BLI RBD-specific serum antibody titers in the 2 and 10 μg S2P-immunized groups were significantly higher than all other groups. ( E ) Correlation between neutralization titer and BLI RBD-specific antibody titer did not exist in the S2P-NDVLP-immunized groups, but a weak correlation existed in the S2P-immunized groups. ( F ) Quality of the immune responses elicited by immunization was calculated as the ratio of the neutralization titer to the total ELISA IgG titer, plotted as the mean and SD. In ( A , B , D ), the Kruskal–Wallis test was used to test differences between the groups, and P-values were designated as *: p < 0.05; **: p < 0.01; ***: p < 0.001; ****: p < 0.0001, and geometric mean titers were marked with a red bar. Spearman correlation was used in ( C , E ).

Article Snippet: A codon-optimized plasmid CMV/R-SARS-CoV-2-encoding D614G spike variant was constructed and cotransfected with a lentivirus backbone, luciferase reporter, and human transmembrane protease serine 2 (TMPRSS2) in HEK293T/17 cells (ATCC #CRL-11268).

Techniques: Enzyme-linked Immunosorbent Assay, Neutralization, Variant Assay