rabbit anti sars cov nucleocapsid antibody  (Sino Biological)

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: ELISA analysis of ß-S 13 EM-SARS-CoV-2 VLPs and ß-S WT EM-CoV-2 VLPs for spike ( A ) and RBD ( B ) proteins. Plates were coated with ß-S WT EM-CoV-2 VLPs or ß-SARS-CoV-2 RBD followed by probing with either anti-Spike or anti-RBD antibody in 2-fold dilutions from 1:400 to 1:51200 (mean ± SD). ELISAs were performed three times and each dilution tested in triplicate. Western immunoblot analysis of ß-S 13 EM-SARS-CoV-2 VLPs compared with ß-S WT EM-CoV-2 VLPs for the presence of ( C ) Spike protein ( D ) Membrane and ( E ) Envelope protein. Amicon ultrafiltration consistently resulted in a higher yield S, M and E proteins than sucrose cushion ultracentrifugation as shown by Western blot analysis of the VLPs ( F ) Negative stain electron microscopy showing morphology of ( F ) ß-S 13 EM-SARS-CoV-2 VLPs and ( G ) ß-S WT EM-CoV-2 VLPs.

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Membrane, Staining, Electron Microscopy

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: ELISA analysis of ß-S 13 EM-SARS-CoV-2 VLPs for spike ( A ) and RBD ( B ) proteins. Plates were coated with ß-S13EM-SARS-CoV-2 VLPs followed by probing with either anti Spike or anti-RBD antibody in 2-fold dilutions from 1:400 to 1:51200 (mean ± SD). ELISAs were repeated three times and each dilution tested in triplicate ( C ) Western immunoblot analysis of ß-S 13 EM-SARS-CoV-2 VLPs showing the presence of spike, membrane and the dimeric form of envelope protein. D Immunoprecipitation of ß-S 13 EM-SARSCoV-2 VLPs with anti-Wu-RBD antibody and probing with anti-Wu-S antibody. Anti-N antibody failed to immunoprecipitate VLPs. ß-S 13 EM-SARS-CoV-2 and Ancestral SARS-CoV-2 VLPs were included as positive controls for S protein.

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Membrane, Immunoprecipitation

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: A , B , C Transmission EMs showing ß-S 13 EM-SARS-CoV-2 VLPs of 50-100 nm and with characteristic spikes. D , E Immunogold EM showing binding of gold beads with anti-ß-RBD antibody to RBDs of spikes on a single VLP and cluster of VLPs. F Colour enhanced image of Immunogold EM highlighting the spacing of anti-ß-RBD coated gold beads to spikes of a ß-S 13 EM-SARS-CoV-2 VLP. G IEM of Delta SARS-CoV-2 virus as a positive control. Scale bar = 100 nm.

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Transmission Assay, Binding Assay, Virus, Positive Control

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: A sVNT assay (with Ancestral RBD-HRP) was performed on immune sera from mice immunised with ß-SARS-CoV-2 VLPs formulated with Addavax, or PBS only and compared to respective prebleed samples. For prebleed samples, owing to small volumes collected, sera from respective groups were pooled and tested in duplicate. Day 28 sera for each mouse in each vaccine group were tested individually in duplicate. Data were analysed using one-way ANOVA and Tukey’s Multiple Comparisons test. B Samples from day 42 were tested in a multiplex sVNT inhibition assay for their ability to inhibit the binding to ACE2 to SARS-CoV-2 Alpha, Beta, Gamma, Delta, Delta + , Lambda, Mu, Omicron BA.1, BA.2, BA.5, XBB.1 and XBB.1.5. Multiplex assays with sera from two separate immunisation series were performed in duplicate. The arithmetic mean titres of the half maximal inhibitory dilution (ID 50 ) are shown for each sample and the range of titres shown in the legend bar.

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Multiplex Assay, Inhibition, Binding Assay

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: A , B C57BL/6 mice were injected with two doses of 10ug ß-S 13 EM-SARS-CoV-2 VLP, i.m., formulated or not with Addavax (50% vol/vol) 14 days apart. Control mice received PBS. Mice were euthanised on day 21 and splenocytes (2 × 10 5 per well) were restimulated in vitro using primary DC from naïve mice (2 × 10 5 per well) pre-cultured with 5 µg/well ß-S 13 EM-SARS-CoV-2 VLP. Numbers of IFN-γ-producing cells were measured using ELISpot. All spleens in each group ( n = 3) were pooled and analysed in triplicate wells per group. Data are from a single experiment and were analysed using one-way ANOVA. A Number of spots/well. B Average total number of responding splenocytes per mouse. Data were log-transformed and compared using One-Way ANOVA and Tukey’s Multiple Comparisons test. Data are from a single experiment with 3 mice/group, and were log-transformed and compared using one-way ANOVA and Tukey’s Multiple Comparisons test. C , D As in ( A , B ), but mice were vaccinated with 20 µg ß-S 13 EM-SARS-CoV-2 VLPs. In this case, 10 5 primary DC were used per well for in vitro restimulation. Data are from a single experiment with three mice/group, and were log-transformed and compared using one-way ANOVA and Tukey’s Multiple Comparisons test. E Mice were vaccinated and splenocytes extracted, restimulated with ß-S 13 EM-SARS-CoV-2 VLPs in vitro, and screened for IFNγ production using ELISpot as in A (10 µg ß-S 13 EM-SARS-CoV-2 VLP dose), but CD4 or CD8 T cells were depleted prior to culture using magnetic beads mixed with anti-CD4 and anti-CD8 antibodies. Non-depleted samples were incubated with beads without antibodies. Data are from a single experiment and were log-transformed and analysed using two-way ANOVA and Tukey’s multiple comparisons test to account for comparisons between the variables of immunisation groups and treatment groups. F Mice were vaccinated and splenocytes extracted, restimulated with ß-S 13 EM-SARS-CoV-2 VLPs in vitro, and screened for IFNγ production using ELISpot as in C (20 µg ß-S 13 EM-SARS-CoV-2 VLP dose), but CD4+ or CD8 + T cells were depleted prior to culture using magnetic beads mixed with anti-CD4 and anti-CD8 antibodies. Non-depleted samples were incubated with beads without antibodies. Data are from a single experiment and were log-transformed and analysed using two-way ANOVA and Tukey’s multiple comparisons test. G Detection of CD8 + T cell responses elicited by the ß-S 13 EM-SARS-CoV-2 vaccine formulated with Addavax. C57BL/6 mice were vaccinated twice with 20 µg ß-S 13 EM-SARS-CoV-2 VLPs as in C. S 539-546 Tetramer staining was used to identify Spike-specific CD8 + T cells among splenocytes. Data are representative of three independent experiments and were log transformed and compared using an unpaired t-test. H Mice were vaccinated twice with 20 µg of ß-S 13 EM-SARS-CoV-2 VLPs as in C, and CD8 T cells were enriched from their spleens, and restimulated in vitro using S 539-546 peptide. ELISpot was performed to detect IFNγ responses. Data are from a single experiment and were log-transformed and analysed using two-way ANOVA and uncorrected Fisher’s Least Significant Difference test (Immunisation schedule figure created with BioRender.com).

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Injection, Control, In Vitro, Cell Culture, Enzyme-linked Immunospot, Transformation Assay, Magnetic Beads, Incubation, Staining

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: Titres of virus in lungs of mice (five per group) vaccinated subcutaneously with two doses 14 days apart with PBS, 20, 10 or 5 μg of ß-SEM-SARS-CoV-2 VLP/Addavax. Mice were aerosol challenged with ß-SARS-CoV-2 virus 28 days after the second immunisation. Age and sex matched PBS injected control C57/BL6 mice ( n = 5) were also challenged. Three days after challenge, mice were euthanized and the titre of infectious virus (TCID 50 /ml) in lungs of individual mice were determined. Challenge experiments were performed twice and titrations repeated in triplicate. Means for each group are depicted. Data were analysed using one-way ANOVA and Tukey’s Multiple Comparisons test. LOD Limit of detection (Immunisation schedule figure created with BioRender.com).

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Virus, Aerosol, Injection, Control

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: A Mice were immunised with 2 doses of 5, 10 or 20 µg of vaccine with or without MF59 2 weeks apart, bled at days 0, 14 and 28. Anti-ß-S 13 EM-SARS-CoV-2 VLP antibody responses were assessed using ELISA. Immunisations were performed twice, with 5 mice in each vaccine group. Sera for each group at respective time points were pooled and tested in triplicate. ELISAs were performed twice. Data from groups of mice were compared using one-way ANOVA and Tukey’s Multiple Comparisons test. Bar indicates mean value; each dot represents one mouse. Geometric mean titre (GMT) values are displayed for each group at day 0, 14 and 28. B Mice were immunised with 2 doses of 5, 10 or 20 µg of vaccine with MF59 2 weeks apart, bled at days 0, 14 and 28. Anti-ß-RBD antibody responses were assessed using ELISA, as before. Immunisations were performed twice with five mice in each vaccine group. Sera for each group at respective time points were pooled and tested in triplicate. ELISAs were performed twice. Data from groups of mice were compared using one-way ANOVA and Tukey’s Multiple Comparisons test. Geometric mean titre (GMT) values are displayed for each group at day 0,14 and 28. C Heat map of sNAb responses against SARS-CoV-2 variants. Samples from day 28 were tested in a multiplex sVNT inhibition assay for their ability to inhibit the binding to ACE2 to SARS-CoV-2 Alpha, Beta, Gamma, Delta, Delta+, Lambda, Mu, Omicron BA.1, BA.2, BA.5, XBB.1 and XBB.1.5. Multiplex assays were performed in duplicate. The mean titres of the half maximal inhibitory dilution (ID 50 ) are shown for each sample and the range of titres shown in the legend bar. (Immunisation schedule figure created with BioRender.com).

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Enzyme-linked Immunosorbent Assay, Multiplex Assay, Inhibition, Binding Assay

Journal: NPJ Vaccines

Article Title: Preclinical development of a cross-protective β-SARS-CoV-2 virus-like particle vaccine adjuvanted with MF59

doi: 10.1038/s41541-025-01355-y

Figure Lengend Snippet: Titres of virus in lungs of mice (five per group) vaccinated subcutaneously with two doses, 14 days apart, of 5, 10 or 20 μg of ß-SEM-SARS-CoV-2 VLP/MF59. To ensure mice could be infected with Omicron BA.5 and to maintain comparability across variant groups, mice were transduced intranasally with AAV-hACE2. Mice were challenged by intranasal inoculation with Beta-, Delta- or Omicron BA.5-SARS-CoV-2 viruses 28 days after the second immunisation. Age and sex-matched PBS vaccinated control C57/BL6 mice ( n = 5) were also challenged. Three days after challenge, mice were euthanised and the titre of infectious virus (TCID 50 ) in lungs of individual mice was determined in triplicate. Immunisation and challenge studies were performed twice. Means for each group are depicted. Data were analysed using one-way ANOVA and Tukey’s Multiple Comparisons test. LOD limit of detection. (Immunisation schedule figure created with BioRender.com).

Article Snippet: Briefly, cells were permeabilized with 0.1% Triton X-100 (v/v) in 0.2% BSA/PBS (w/v) for 10 min, blocked in 0.2% BSA/PBS for 10 min and labelled with 1 μg/mL of rabbit anti-SARS-CoV nucleocapsid antibody (Sinobiological #40143-R004) for 1 h, all at room temperature.

Techniques: Virus, Infection, Variant Assay, Control