Journal: Phytomedicine

Article Title: Kaempferol inhibits SARS-CoV-2 invasion by impairing heptad repeats-mediated viral fusion

doi: 10.1016/j.phymed.2023.154942

Figure Lengend Snippet: Kae prevents the invasion of SARS-CoV-2 particles into host cells. (A and B), immunofluorescence staining (A) or enzyme-linked immunosorbent assay (ELISA) analysis (B) of SARS-CoV-2 virus-like particles (VLPs) invasion into human induced pluripotent stem cells (hiPSC)-derived alveolar epithelial cells type II (AECII). Control: dimethyl sulfoxide (DMSO)-treated cells; spike (S) protein neutralizing antibody (S NAb): positive control; and Kae: 10 µM in DMSO.(C and D), the entry of SARS-CoV-2 VLPs into lungs of hACE2 transgenic mice evaluated by immunofluorescence staining (C) or luciferase activity (D).Control: 0.5% carboxymethylcellulose sodium-treated mice; ursodeoxycholic acid (UDCA): positive control; and Kae: indicated dosages in 0.5% carboxymethylcellulose sodium. (A and C), VLPs, ACE2, F-actin and nucleus are shown as indicated colors. Scale bar = 25 µm. (B), ***, p < 0.005 vs Blank. #, p < 0.05 and ##, p < 0.01 vs Control. (D), n = 4, ***, p < 0.005 vs Blank. ##, p < 0.01; and ###, p < 0.005 vs Control. ns: no statistical differences vs Control.

Article Snippet: The procedure involved incubating 100 ng/ml of human Fc-tagged SARS-CoV-2 S-RBD protein (40592-V02H, Sino Biological) with either 2‰ DMSO or 10 µM Kae for 30 min, then adding the mixture to HEK-293F-ACE2 cells and incubating for an additional 30 min at 37˚C.

Techniques: Immunofluorescence, Staining, Enzyme-linked Immunosorbent Assay, Virus, Derivative Assay, Control, Positive Control, Transgenic Assay, Luciferase, Activity Assay

Journal: Phytomedicine

Article Title: Kaempferol inhibits SARS-CoV-2 invasion by impairing heptad repeats-mediated viral fusion

doi: 10.1016/j.phymed.2023.154942

Figure Lengend Snippet: Kae targets coronaviral fusion. (A), flow cytometry analyses of the binding affinity between spike (S) protein receptor-binding domain (S-RBD) and HEK-293F cells expressing angiotensin-converting enzyme 2 (ACE2) in the presence or absence of Kae with the S protein neutralizing antibody (S NAb) as a positive control. ***, p < 0.005 vs Blank. ###, p < 0.005, and ns, no statistical differences vs Control. (B), impacts of Kae on SARS-CoV-2 endocytosis into Vero E6 cells with S NAb as a positive control. ***, p < 0.005, and ns, no statistical differences vs Control. (C, D and E), effects of Kae on membrane fusion mediated by SARS-CoV-2 variants (C), or SARS-CoV (D), or MERS-CoV (E) with transmembrane serine protease 2 (TMPRSS2) inhibitor, camostat mesylate (CM), as a positive control. Scale bar = 300 µm. **, p < 0.01; and ***, p < 0.005 vs Control. Control: dimethyl sulfoxide (DMSO)-treated cells; DPP4: dipeptidyl peptidase 4; ns: no statistical differences vs Control.

Article Snippet: The procedure involved incubating 100 ng/ml of human Fc-tagged SARS-CoV-2 S-RBD protein (40592-V02H, Sino Biological) with either 2‰ DMSO or 10 µM Kae for 30 min, then adding the mixture to HEK-293F-ACE2 cells and incubating for an additional 30 min at 37˚C.

Techniques: Flow Cytometry, Binding Assay, Expressing, Positive Control, Control, Membrane

Journal: Phytomedicine

Article Title: Kaempferol inhibits SARS-CoV-2 invasion by impairing heptad repeats-mediated viral fusion

doi: 10.1016/j.phymed.2023.154942

Figure Lengend Snippet: Kae interferes with the S2 subunit of SARS-CoV-2 spike protein. (A), immunoblots representing the cleavage and abundance of SARS-CoV-2 spike (S) protein induced by Kae, with camostat mesylate (CM) as a positive control. (B), thermal stability determinations for full-length S protein, S1 and S2 subunits of SARS-CoV-2 in the presence or absence of Kae. Control: dimethyl sulfoxide (DMSO)-treated cells.

Article Snippet: The procedure involved incubating 100 ng/ml of human Fc-tagged SARS-CoV-2 S-RBD protein (40592-V02H, Sino Biological) with either 2‰ DMSO or 10 µM Kae for 30 min, then adding the mixture to HEK-293F-ACE2 cells and incubating for an additional 30 min at 37˚C.

Techniques: Western Blot, Positive Control, Control

Journal: Phytomedicine

Article Title: Kaempferol inhibits SARS-CoV-2 invasion by impairing heptad repeats-mediated viral fusion

doi: 10.1016/j.phymed.2023.154942

Figure Lengend Snippet: Kae impairs HR1 and HR2 of SARS-CoV-2 S2 subunit. (A and B), circular dichroism (CD) spectra analyses of synthesized heptad repeat 1 (HR1) and heptad repeat 2 (HR2) peptide, the formation of six-helix bundle (6-HB, HR1+HR2), and impacts of Kae on 6-HB, HR1 and HR2 using isopropanol as solvent controls. (C, D, E and F), native polyacrylamide gel electrophoresis (N-PAGE) assessment of effects of Kae on HR1, HR2 and 6-HB. *, p < 0.05; **, p < 0.01; and ***, p < 0.005 vs Control. ns: no statistical differences vs Blank. (E and F), the stability of wildtype (WT) and mutant (MT) HR2 in the absence (E) or presence (F) of Kae compared to corresponding wildtype peptide. (G), atomic force microscopy (AFM) topographic images before and after adding Kae to HR1, HR2 and 6-HB. Scale bar = 4 µm. (C, D, F and G), Control: dimethyl sulfoxide (DMSO)-treated peptides.

Article Snippet: The procedure involved incubating 100 ng/ml of human Fc-tagged SARS-CoV-2 S-RBD protein (40592-V02H, Sino Biological) with either 2‰ DMSO or 10 µM Kae for 30 min, then adding the mixture to HEK-293F-ACE2 cells and incubating for an additional 30 min at 37˚C.

Techniques: Circular Dichroism, Synthesized, Solvent, Polyacrylamide Gel Electrophoresis, Control, Mutagenesis, Microscopy

Journal: Phytomedicine

Article Title: Kaempferol inhibits SARS-CoV-2 invasion by impairing heptad repeats-mediated viral fusion

doi: 10.1016/j.phymed.2023.154942

Figure Lengend Snippet: Schematic diagram of the inhibitory mechanism of Kae against SARS-CoV-2 invasion. Instead of blocking the interaction between angiotensin-converting enzyme 2 (ACE2) and spike (S) protein receptor-binding domain (S-RBD), Kae disrupts heptad repeat 1 (HR1) and heptad repeat 2 (HR2) segments in the S2 subunit of SARS-CoV-2 S protein, leading to inhibitions of viral fusion-mediated invasion of SARS-CoV-2.

Article Snippet: The procedure involved incubating 100 ng/ml of human Fc-tagged SARS-CoV-2 S-RBD protein (40592-V02H, Sino Biological) with either 2‰ DMSO or 10 µM Kae for 30 min, then adding the mixture to HEK-293F-ACE2 cells and incubating for an additional 30 min at 37˚C.

Techniques: Blocking Assay, Binding Assay

Journal: Frontiers in Immunology

Article Title: Spike-Dependent Opsonization Indicates Both Dose-Dependent Inhibition of Phagocytosis and That Non-Neutralizing Antibodies Can Confer Protection to SARS-CoV-2

doi: 10.3389/fimmu.2021.808932

Figure Lengend Snippet: Neutralization assays identify one monoclonal as blocking the ACE2-Spike protein interaction. (A) SPR analysis of the binding of monoclonal antibodies to the RBD domain of the Spike protein. PBS served as a negative control, and the intact RBD was our positive control for ACE2 binding. (B) The 96 antibodies which we produced were assayed for neutralization potential in a Spike-bead-based neutralization assay. Spike-beads [such as the ones used in (B) ] were opsonized with the antibodies in 96 well plates. The beads were then centrifuged, reconstituted in fresh media, and added to HEK293-ACE2 cells at a ratio of 20 beads per cell and imaged with automated microscopy. The data is from 4 pooled experiments and is presented as bead association normalized percentage. Error bars indicate the SEM for the replicate experiments. (C) The 10 Spike-ELISA reactive antibodies were assayed for pseudovirus neutralization. A firefly luciferase encoding pseudotype lentivirus was used to infect HEK239-ACE2 cells. Antibody serial dilutions were used to block the viral entry into the HEK293-ACE2 cells. Nonlinear regression lines were fitted for the three antibodies that showed a higher than 50% reduction of infectivity. Those antibodies were highlighted in green (Ab57), blue (Ab36), and red (Ab59). Created with BioRender.com .

Article Snippet: The antigens, produced in HEK293 cells, were obtained from SinoBiological (Beijing, China; product numbers: SARS-CoV-2 Spike RBD: 40592-V08H; SARS-CoV-2 Spike RBD-N501Y: 40592-V08H82; SARS-CoV-2 Spike RBD-E484K: 40592-V08H84; SARS-CoV-2 Spike RBD-K417N, E484K, N501Y: 40592-V08H85; SARS-CoV-2 Spike S1 HV69-70 deletion, Y144 deletion, N501Y, A570D, D614G, P681H: 40591-V08H12).

Techniques: Neutralization, Blocking Assay, Binding Assay, Negative Control, Positive Control, Produced, Microscopy, Enzyme-linked Immunosorbent Assay, Luciferase, Infection

Journal: Cell reports

Article Title: Beta-spike-containing boosters induce robust and functional antibody responses to SARS-CoV-2 in macaques primed with distinct vaccines

doi: 10.1016/j.celrep.2023.113292

Figure Lengend Snippet: A subunit booster improves antibody profiles in both macaques primed with two-dose mRNA vaccines and macaques primed with two-dose subunit vaccines (A) Schematic representation of the cohorts: in the mRNA-primed cohort, 16 macaques were evenly split into four groups, each group was primed with mRNA vaccines in different modalities on days 0 and 21 and boosted with a subunit booster on day 210, and blood samples were collected 2 weeks after the second primary series on day 35 and on days 205 and 224. In the protein-primed cohort, 24 macaques were split into five groups, each group was primed with subunit vaccines in different modalities on days 0 and 21 and boosted with a subunit booster on day 202, and blood samples were collected 2 weeks after the second primary series vaccination on day 35 and on days 196 and 216. (B) The dot plots show the SARS-CoV-2 ancestral spike-specific IgG1 titer and the ability of the spike-specific antibodies to bind to the low-affinity Fcγ receptors (FcγRIIA and FcγRIIIA) across the macaques primed with mRNA vaccines and boosted with a subunit booster in mRNA prime (top) at different time points: peak (day 35), pre-boost (day 205), and post-boost (day 224), and the macaques primed with subunit vaccines and boosted with a subunit booster in protein prime (bottom) at different time points: peak (day 35), pre-boost (day 205), and post-boost (day 224). Each dot represents a different macaque. Each bar represents the median of each group. (C) The dot plots show antibody-dependent cellular phagocytosis (ADCP), antibody-dependent neutrophil phagocytosis (ADNP), antibody-dependent complement deposition (ADCD), and antibody-dependent natural killer (NK) cell activity (ADNK), as measured by CD107a degranulation, against SARS-CoV-2 ancestral spike in macaques primed with mRNA vaccines and boosted with a subunit booster in mRNA prime (top) at different time points: peak (day 35), pre-boost (day 205), and post-boost (day 224), and the macaques primed with subunit vaccines and boosted with a subunit booster in protein prime (bottom) at different time points: peak (day 35), pre-boost (day 196), and post-boost (day 216). *p < 0.05 against the no-AS03 post-boost group, ns, not significant.

Article Snippet: SARS-CoV-2 Gamma RBD , Sino Biological , 40592-V08H86.

Techniques: Vaccines, Activity Assay

Journal: Cell reports

Article Title: Beta-spike-containing boosters induce robust and functional antibody responses to SARS-CoV-2 in macaques primed with distinct vaccines

doi: 10.1016/j.celrep.2023.113292

Figure Lengend Snippet: IgA toward SARS-CoV-2 spikes is expanded by Beta-containing component boosters in protein-primed NHPs (A) Mean IgA binding to indicated SARS-CoV-2 VOCs in mRNA-primed NHPs. Shown are the values after the primary series, pre-boost, and post-boost against the indicated VOC spike or tetanus as a negative control. Shown on the y axis is the IgA mean fluorescent intensity (MFI) of binding to the antigen, and on the right is the color scheme. (B) Same as (A) but for NHPs who received a protein primary vaccines series. Statistical significance (Mann-Whitney U test) of peak responses after primary series and post-boost was determined to show expansions of IgA binding breadth. *p < 0.05, **p < 0.01, and ***p < 0.001.

Article Snippet: SARS-CoV-2 Gamma RBD , Sino Biological , 40592-V08H86.

Techniques: Binding Assay, Negative Control, Vaccines, MANN-WHITNEY

Journal: Cell reports

Article Title: Beta-spike-containing boosters induce robust and functional antibody responses to SARS-CoV-2 in macaques primed with distinct vaccines

doi: 10.1016/j.celrep.2023.113292

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: SARS-CoV-2 Gamma RBD , Sino Biological , 40592-V08H86.

Techniques: Recombinant, Virus, Software, Labeling, Luminex

Journal: iScience

Article Title: Placental transfer dynamics and durability of maternal COVID-19 vaccine-induced antibodies in infants

doi: 10.1016/j.isci.2024.109273

Figure Lengend Snippet: Maternal vaccine-induced SARS-CoV-2 antibodies persist in 2-month and 6-month infants Dot plots showing (A) Spike-specific WT (top) and RBD-specific WT antibody levels and (B) FcR profiles of maternal (M), cord (C), 2-month-old infants (2 m), and 6-month-old infants (6 m). Significance was determined by a Kruskal-Wallis test followed by Benjamini-Hochberg procedure for multiple hypotheses correction. If statistically significant, a two-sided Mann-Whitney U test was performed. (∗p < 0.05, ∗∗p < 0.01,∗∗∗p < 0.001,∗∗∗∗p < 0.0001). The dashed line represents the level of detection for each feature, defined as one standard deviation above the mean value for assay negative biological controls (SARS-CoV-2 negative, unvaccinated samples).

Article Snippet: SARS-CoV-2 RBD delta , Sino Biological , CAT # 40592-V08H115.

Techniques: MANN-WHITNEY, Standard Deviation

Journal: iScience

Article Title: Placental transfer dynamics and durability of maternal COVID-19 vaccine-induced antibodies in infants

doi: 10.1016/j.isci.2024.109273

Figure Lengend Snippet: Maternal vaccine timing drives IgG levels and Fc-receptor binding profiles in 2-month infants Infants whose mothers were vaccinated in the late second and early third trimester (received at least one dose of mRNA vaccine after 28 weeks) had the highest levels of antibodies at 2 months. (A) Diagram showing the grouping strategy for 2-month-old infants based on the dose 1 (D1) and dose 2 (D2) COVID-19 vaccination. Group 1: N = 12 , Group 2: N = 11, Group 3: N = 10, Group 4: N = 6, Group 5: N = 8. (B) Univariate plots showing Spike-specific WT antibody and Fc-receptor (FcR) levels of each 2-month-old infant group (as shown in in A) Significance was determined by a Kruskal-Wallis test followed by Benjamini-Hochberg procedure for multiple hypotheses correction. If statistically significant, a two-sided Mann-Whitney U test was performed. (∗p < 0.05, ∗∗∗∗p < 0.0001). The middle line represents the median, whereas the other lines represent the first and third quartiles. The dashed line represents the level of detection for each feature, defined as one standard deviation above the mean value for assay negative biological controls (SARS-CoV-2 negative, unvaccinated samples). (C) (Top) A partial-least squares discriminant model (PLSDA) was built using LASSO feature regularization and variable selection of the infant cohort group, resulting a reduced set of features that were consistently in at least 80 of the 100 LASSO models. (Bottom) Variable importance in projection (VIP) score of the selected features. The magnitude indicates the importance of the features in driving the separation in the model. The color of the feature corresponds to the group to which the feature was enriched.

Article Snippet: SARS-CoV-2 RBD delta , Sino Biological , CAT # 40592-V08H115.

Techniques: Binding Assay, MANN-WHITNEY, Standard Deviation, Selection

Journal: iScience

Article Title: Placental transfer dynamics and durability of maternal COVID-19 vaccine-induced antibodies in infants

doi: 10.1016/j.isci.2024.109273

Figure Lengend Snippet: Maternal vaccine administration at >32 weeks is associated with reduced antibody persistence in 6-month infants (A) The polar plots depict the mean percentile ranks of the antibody features within 6 months infants whose mothers were vaccinated with the first dose of the mRNA COVID-19 vaccine <20 weeks ( N = 20), 20–28 weeks ( N = 26), and >28 weeks ( N = 30). Each wedge represents an antibody feature, and the size of the wedge depicts the mean percentile ranging from 0 to 1. (B) Linear regression models fit the relationship between gestation age of dose 1 (days) (those vaccinated between 28 and 32 weeks) and anti-RBD WT IgG1, anti-S1 FcγR2AR, and anti-RBD Gamma FcγR3B. The rho and p values are reported in the plot. The dashed line represents the level of detection for each feature, defined as one standard deviation above the mean value for assay negative biological controls (SARS-CoV-2 negative, unvaccinated samples). (C) Linear regression models fit the relationship between gestation age of dose 1 (days) (those vaccinated between 32 and 36 weeks) and anti-RBD WT IgG1, anti-S1 FcγR2AR, and anti-RBD Gamma FcγR3B. The rho and p values are reported in the plot. The dashed line represents the level of detection for each feature, defined as one standard deviation above the mean value for assay negative biological controls (SARS-CoV-2 negative, unvaccinated samples). (D) The heatmap shows the Spearman correlation score for each feature against gestational age (days) within infant samples whose mothers received the first dose of the mRNA COVID-19 vaccine between 28 and 32 weeks or 32–36 weeks. The red boxes indicate samples with a positive Spearman correlation and blue boxes indicate samples with a negative Spearman correlation. Within each group, gray boxes are features which were excluded from the analysis (if >70% of values fell below the threshold of detection, defined as less than one standard deviation above the mean of SARS-CoV-2 negative, unvaccinated control samples).

Article Snippet: SARS-CoV-2 RBD delta , Sino Biological , CAT # 40592-V08H115.

Techniques: Standard Deviation

Journal: iScience

Article Title: Placental transfer dynamics and durability of maternal COVID-19 vaccine-induced antibodies in infants

doi: 10.1016/j.isci.2024.109273

Figure Lengend Snippet: IgG subclasses and Fc-receptor binding capacity of antibodies against SARS-CoV-2 variants of concern in umbilical cord blood, 2-month, and 6-month infants (A) The heatmap shows the ratio of infants with detectable antibody titer or FcR binding levels N = 76 for cord (C), N = 55 for 2-month-old infants (2 M), and N = 80 for 6-month-old infants (6 M) against the Spike or RBD region of wild-type (WT) strain and variants of concern (VOCs) including Alpha, Beta, Delta, Gamma, and Omicron. (B) Univariate plots showing RBD-specific Omicron antibody IgG levels (top) and FcR binding profiles in cord (C), 2-month-old infants (2 m), and 6-month-old infants (6 m). Significance was determined by a Kruskal-Wallis test followed by Benjamini-Hochberg procedure for multiple hypotheses correction. If statistically significant, a two-sided Mann-Whitney U test was performed. (∗p < 0.05, ∗∗p < 0.01,∗∗∗p < 0.001,∗∗∗∗p < 0.0001). The dashed line represents the level of detection for each feature, defined as one standard deviation above the mean value for assay negative biological controls (SARS-CoV-2 negative, unvaccinated samples).

Article Snippet: SARS-CoV-2 RBD delta , Sino Biological , CAT # 40592-V08H115.

Techniques: Binding Assay, MANN-WHITNEY, Standard Deviation

Journal: iScience

Article Title: Placental transfer dynamics and durability of maternal COVID-19 vaccine-induced antibodies in infants

doi: 10.1016/j.isci.2024.109273

Figure Lengend Snippet:

Article Snippet: SARS-CoV-2 RBD delta , Sino Biological , CAT # 40592-V08H115.

Techniques: Recombinant, Produced, Generated, Software, Luminex