Journal: NPJ Vaccines

Article Title: Bivalent norovirus mRNA vaccine elicits cellular and humoral responses protecting human enteroids from GII.4 infection

doi: 10.1038/s41541-024-00976-z

Figure Lengend Snippet: Expi293F cells were transfected with mRNA-encoding ( A ) norovirus GI.1 or ( B ) GII.4 VP1 and analyzed for protein expression by western blot as described in Methods. The supernatant was collected, clarified, ultrafiltered, and visualized via NSEM. Representative micrographs are shown from cells transfected with ( C ) GI.1 or ( D ) GII.4, with n = 5 micrograph images per group. E Experimental schematic of the study. Balb/c mice ( n = 10 per group) were immunized on days 0 and 28 with 10 μg of mRNA-LNP vaccine encoding Norwalk1968/GI.1 or CapeTown2012/GII.4 VP1, or empty LNP as control at an equivalent dose. Sera were collected at various time points (4, 8, 14, 22, 26, and 34 weeks post-prime) and screened against ( F ) Norwalk1968/GI.1 and ( G ) CapeTown2012/GII.4 VLPs to assess their ability to block VLP binding to its carbohydrate ligand in a surrogate neutralization assay. The data was analyzed using GraphPad Prism, and the dilution at which 50% of VLP-ligand binding was blocked (ID50) was calculated. Sera that did not block at least 50% of VLP-ligand binding were assigned a titer of 0.5X the lower limit detection (ID50 = 25) and marked below this limit (dashed line). The nAb titer is presented as 50% inhibitory dilution (ID50) values (Mean ± SD), n = 10 per group (5 male and 5 female). The nAb titers were compared with the values from Day 56 for each strain, ∗ p < 0.05 F:(D56 vs D28 p = 0.0135) G:(D56 vs D28 p = <0.0001, D56 vs D150 p = <0.0001, D56 vs D180 p = <0.0001, d56 vs d240 p = <0.0001), one-way ANOVA, Tukey’s post hoc tests. A red square symbol indicates nAb titers in a serum sample collected on Day 14 from a patient infected with the GII.4 norovirus strain. nAb titer values for empty LNP are not shown because they were below the level of detection. Schematic for Fig. 1E created with Biorender.

Article Snippet: Splenocytes were stimulated with 80 μl of a synthetic peptide pool of CapeTown/GII.4 VP1 (150 peptides, each with 15 AA length and 11 AA overlap, a purity level between 80 and 90%, synthesized by GenScript) at a final concentration of 1.5 μg/mL, and 1 μg/mL of CD28 (co-stimulatory signal) for 1 h at 37 °C.

Techniques: Transfection, Expressing, Western Blot, Control, Blocking Assay, Binding Assay, Neutralization, Ligand Binding Assay, Infection

Journal: NPJ Vaccines

Article Title: Bivalent norovirus mRNA vaccine elicits cellular and humoral responses protecting human enteroids from GII.4 infection

doi: 10.1038/s41541-024-00976-z

Figure Lengend Snippet: Balb/c mice ( n = 10 per group) were immunized on days 0 and 28 with 10 μg of mRNA-LNP vaccine encoding Norwalk1968/GI.1 or CapeTown2012/GII.4 VP1, or empty LNP as control at an equivalent dose. A nAb response titers to three genogroup I strains (GI.1, GI.3, and GI.5) and ( B ) against four genogroup II strains (GII.3, GII.4 Sydney 2012, GII.4 New Orleans 2009, and GII.4 Farmington Hills 2002) at day 56 post prime in a surrogate neutralization assay. The nAb titer is presented as 50% inhibitory dilution (ID50) values (Mean ± SD), n = 10 per group (5 male and 5 female). Only the cross-reactivity nAb titer values of GII.4 variants were compared, ∗ p < 0.05, (2012 vs 2009 p = <0.0001, 2012 vs 2002 p = <0.0001) one way-ANOVA, Tukey’s post hoc tests.

Article Snippet: Splenocytes were stimulated with 80 μl of a synthetic peptide pool of CapeTown/GII.4 VP1 (150 peptides, each with 15 AA length and 11 AA overlap, a purity level between 80 and 90%, synthesized by GenScript) at a final concentration of 1.5 μg/mL, and 1 μg/mL of CD28 (co-stimulatory signal) for 1 h at 37 °C.

Techniques: Control, Neutralization

Journal: NPJ Vaccines

Article Title: Bivalent norovirus mRNA vaccine elicits cellular and humoral responses protecting human enteroids from GII.4 infection

doi: 10.1038/s41541-024-00976-z

Figure Lengend Snippet: A Experimental schematic of the study. Balb/c and C57BL/6 mice were i.m. immunized on days 0 and 28 with varying doses (0.25 μg, 0.5 μg, 1 μg, and 3 μg) of each mRNA-LNP vaccine encoding Norwalk1968/GI.1 and CapeTown2012/GII.4 VP1, or empty LNP as control. Sera were collected 4 weeks after the prime and after the boost, and then screened against ( B , D ) Norwalk1968/GI.1 and ( C , E ) Sydney2012/GII.4 VLP to evaluate their capacity to block VLP binding to their carbohydrate ligand in a surrogate neutralization assay. The nAb titer is presented as 50% inhibitory dilution (ID50) values (Mean ± SEM), n = 5. Dashed line = limit of detection. nAb titer values for empty LNP are not shown because they were below the level of detection. Only nAb titer values within Day 56 were compared, ∗ p < 0.05, B:(D56 0.25 μg vs D56 3 μg p = 0.0007), C:(D56 0.25 μg vs D56 3 μg p = 0.0001) ANOVA, Tukey’s post hoc tests. Schematic for Fig. 3A created with Biorender.

Article Snippet: Splenocytes were stimulated with 80 μl of a synthetic peptide pool of CapeTown/GII.4 VP1 (150 peptides, each with 15 AA length and 11 AA overlap, a purity level between 80 and 90%, synthesized by GenScript) at a final concentration of 1.5 μg/mL, and 1 μg/mL of CD28 (co-stimulatory signal) for 1 h at 37 °C.

Techniques: Control, Blocking Assay, Binding Assay, Neutralization

Journal: NPJ Vaccines

Article Title: Bivalent norovirus mRNA vaccine elicits cellular and humoral responses protecting human enteroids from GII.4 infection

doi: 10.1038/s41541-024-00976-z

Figure Lengend Snippet: A Experimental schematic of the study. Human intestinal enteroids were infected for 2 h at 37 °C with GII.4 Sydney virus (stool sample #20942) that was pre-incubated with serial dilution of sera from mice immunized by i.m. with ( B ) 10 μg or ( C ) 0.25 μg of each mRNA-LNP vaccine encoding Norwalk1968/GI.1 and CapeTown2012/GII.4 VP1 for 1 h at room temperature. Following infection, 3D HIE were washed and cultured in BME for 3 days post-infection (see Methods and Materials). Viral titers were measured by RT-qPCR. * p < 0.05 vs control (empty LNP). B , C Each dot in the graph at a given dilution represents an independent infection performed on a separate day, while each set of bars, i.e., dilutions 1:100 to 1:30,000 ( B ), or 1:300 to 1:10,000 ( C ) represents the dilution series from an individual serum sample. Schematic for Fig. 5A created with Biorender.

Article Snippet: Splenocytes were stimulated with 80 μl of a synthetic peptide pool of CapeTown/GII.4 VP1 (150 peptides, each with 15 AA length and 11 AA overlap, a purity level between 80 and 90%, synthesized by GenScript) at a final concentration of 1.5 μg/mL, and 1 μg/mL of CD28 (co-stimulatory signal) for 1 h at 37 °C.

Techniques: Infection, Virus, Incubation, Serial Dilution, Cell Culture, Quantitative RT-PCR, Control

Journal: Cell host & microbe

Article Title: NON-NEUTRALIZING ANTIBODIES FROM A MARBURG INFECTION SURVIVOR MEDIATE PROTECTION BY FC-EFFECTOR FUNCTIONS AND ENHANCING EFFICACY OF OTHER ANTIBODIES

doi: 10.1016/j.chom.2020.03.025

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: 168 overlapping 15 residue peptides corresponding to MARV strain 200501379 Angola GP sequence were produced by JPT Peptide Technologies GmbH (Berlin, Germany).

Techniques: Recombinant, Infection, Staining, Expressing, Sequencing, Plasmid Preparation, Mutagenesis, Ligation, Peptide Microarray, Software, Flow Cytometry, High Throughput Screening Assay, Cytometry, Chromatography, Microscopy, Magnetic Beads