Enzyme-linked Immunosorbent Assay:
Binding Assay:
Labeling:
Recombinant:
Purification:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Expressing:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Sequencing:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Virus:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Mutagenesis:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Negative Control:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Incubation:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
Concentration Assay:Article Title: Recombinant severe respiratory syndrome coronavirus 2 Rbd trimer protein vaccine capable of generating broad-spectrum cross neutralization activity, and preparation method and use thereof
Article Snippet: .. The purified trimer protein A, protein B and protein C, a trimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.18, which are formed by sequentially connecting three amino acid fragments shown in SEQ ID NO.1), a dimer protein (obtained by performing 293FT cell or CHO cell recombinant expression and chromatographic purification on proteins, with the amino acid sequence shown in SEQ ID NO.19, which are formed by sequentially connecting two amino acid fragments shown in SEQ ID NO.1), an RBD protein (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08B), an RBD protein (K417N, E484K and N501Y; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Beta (B.1.351) strain, and an RBD protein (L452R and E484Q; manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-V08H85) consistent with the virus mutation sites of Kappa (B.1.617.1) strain were diluted with coating liquid to 4 μg/ml, 2 μg/ml, 1 μg/ml, 0.5 μg/ml, 0.25 μg/ml, 0.125 μg/ml, 0.0625 μg/ml, 0.03125 μg/ml, 0.015625 μg/ml, 0.007813 μg/ml, 0.003906 μg/ml, 0.001953 μg/ml, and were coated to a 96-well ELISA plate by 100 μl/well at 4° C. for 8 to 12 h. and a blank well served as negative control; after the plate was washed with a PBST solution, a confining liquid was added for confining at 37° C. for 3 h; after the plate was washed by the PBST solution, a diluted MM43 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40591-MM43; dilutability: 2000 folds) or a diluted MM57 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-MM57; dilutability: 2000 folds) or a diluted R001 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R001; dilutability: 2000 folds) or a diluted R117 monoclonal antibody (manufacturer: Beijing Sino Biological Ltd. Co.; article number: 40592-R117; dilutability: 2000 folds) were respectively added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, a diluted horse radish peroxidase-labeled goat anti-mouse or goat anti-rabbit antibody was added in 100 μl/well for incubation at 37° C. for 1 h; after the plate was washed with the PBST solution, chromogenic solutions A and B were added sequentially for developing for 5 to 10 min at room temperature, and a stop solution C was added; and values were read from a microplate reader at double wavelengths (OD450 nm and 630 nm) to determine a cutoff value, and a curve of protein concentration-absorbance value was drawn. ..
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