Journal: Bioscience Reports

Article Title: Targeting intra-viral conserved nucleocapsid (N) proteins as novel vaccines against SARS-CoVs

doi: 10.1042/BSR20211491

Figure Lengend Snippet: ( A ) N-protein alignment from 4 different Coronaviruses (CoVs): 1. Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2); 2. Severe acute respiratory syndrome coronavirus (SARS-CoV); 3. BAT severe acute respiratory (BAT SARS); 4. BAT coronavirus (BAT-CoV). Yellow color indicates the similarity of sequences among four viruses. The location of Peptide #1, #2, #3 (30 amino acid sequences) is also mentioned. ( B–E ) The whole N-protein vaccination was repeated four times (2-week interval). Red arrow indicates the time points for vaccination. Blood samples were taken before vaccination followed by every 2 weeks until 22nd week. Serum antibodies were detected by using anti-IgM, -IgG1, -IgG2 and anti IgG (H+L) horseradish peroxidase (HRP) conjugated antibodies. Antibody responses can be detected after 2nd vaccination and sustained till last sample collection in mouse#1 (B), #2 (C), #3 (D) and #4 (E). ( F ) Mean data of antibody productions in the BALB/c mice ( n =4). ( G ) Mean antibody production of N-protein vaccination in FVB mice ( n =3). Data represent mean ± S.D.

Article Snippet: Anti N-protein IgG (whole IgG) were detected by goat anti-mouse IgG-HRP (H+L) secondary antibody (Invitrogen 31430).

Techniques:

Journal: Bioscience Reports

Article Title: Targeting intra-viral conserved nucleocapsid (N) proteins as novel vaccines against SARS-CoVs

doi: 10.1042/BSR20211491

Figure Lengend Snippet: ( A ) Peptide sequences were selected based on N-protein conserved regions highlighted in yellow ( A). ( B ) Binding of anti N-polyclonal Abs serum and peptides were tested by ELISA. Whole N-protein was used as control. Peptides, #1, #2, #3 and N-protein were coated respectively with 5 ng and 20 ng/well. Anti-N polyclonal Abs serum (from A mouse at 1:1000 and 1:2000 dilutions) was incubated in each well followed by washing, and incubation with secondary anti-mouse IgG (H+L) (HRP). The optical density (OD) was measured. Anti-N polyclonal Abs bind not only whole N-protein but also enriched binding to Peptide #3, the highest OD compared with Peptide #1 and #2. ( C ) BALB/c mice were immunized with Peptide#3 in 2-week interval, 3 repeats. Red arrow indicates each vaccine time point. Antibody subtype in Anti-Peptide #3 Ab serum were classified by using anti-IgM, -IgG1, -IgG2a and -IgG (H+L) horseradish peroxidase (HRP) conjugated antibodies. Data represents mean ± S. D, n =3. ( D ) Comparison osf antibody subtype induction in N-protein vaccine and Peptide #3 vaccine after vaccination for three times. ( E ) Anti-Peptide #3 Ab serum (from C) were tested by Elisa for the binding capacity. Anti-Peptide #3 Ab serum binds to Peptide #3 and whole N-protein, detected by anti-mouse IgG (H+L) (HRP).

Article Snippet: Anti N-protein IgG (whole IgG) were detected by goat anti-mouse IgG-HRP (H+L) secondary antibody (Invitrogen 31430).

Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Incubation

Journal: Bioscience Reports

Article Title: Targeting intra-viral conserved nucleocapsid (N) proteins as novel vaccines against SARS-CoVs

doi: 10.1042/BSR20211491

Figure Lengend Snippet: ELISA was done to analyze the binding affinity of peptides and N-protein (SARS-CoV2) to in house produced mouse anti SARS-CoV Ab (clone 6H3) . ELISA plate was coated with 5 ng and 20 ng/ well of different peptides and N-protein (SARS-CoV2). Mouse 6H3 antibodies were diluted at 1:1000 and 1:5000. The binding of antibody was detected by anti-mouse IgG (H+L) (HRP). The optical density (OD) was measured.

Article Snippet: Anti N-protein IgG (whole IgG) were detected by goat anti-mouse IgG-HRP (H+L) secondary antibody (Invitrogen 31430).

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay, Produced