histagged pr8 nuclear protein  (Sino Biological)

Journal: bioRxiv

Article Title: Divergent effects of a Treg selective IL-2 mutein on Influenza specific T cell responses

doi: 10.1101/2025.06.10.658962

Figure Lengend Snippet: Mice were infected with X31 and treated with vehicle (blue), or with Fc.Mut24 either a day before (red) or 4 days after (green) infection.. A) Mice were lethally challenged with 5000 FU of PR8 at 30 days post X31 infection. Disease severity based on weight loss from start of lethal challenge in mice treated with Fc.Mut24 one day before (left) or 4 days after (right) initial X31 infection (left) compared with vehicle-treated and X31-naïve mice. B), C) Representative flow cytometry analysis of CD44 expression and NP 366-374 tetramer staining by gated CD8 + T cells in the lungs of vehicle-and Fc.Mut24-tretted mice, and quantification of the number and frequency of CD44 hi Tetramer - T cells in the lungs at 170dpi in mice treated one day prior to B) or 4 days after C) initial X31infection. D), E) X31-immune mice treated with vehicle or Fc.Mut24 were lethally challenged with 5000 FU of PR8 at 170dpi. Disease severity based on weight loss from start of lethal challenge (left). Quantification of Flu-sp CD8 T cells in lung (right) in mice treated with Fc.Mut24 one day prior to D) or 4 days after E) initial X31 infection. *p < 0.05, **p < 0.01, ****p < 0.0001, by unpaired T test and two way ANOVA with mixed effects for weight loss curves in panels A, C, D and E.

Article Snippet: Plates were then washed three times with 100 μL PBS containing 0.1% Tween-20 (PBS-T), followed by incubation with His-tagged PR8 nuclear protein (SinoBiological, Cat# 11675-V08B) at 1:100 dilution in PBS-T (50 μL/well) for 1 hour at 37°C.

Techniques: Infection, Flow Cytometry, Expressing, Staining

Journal: Nature Immunology

Article Title: Influenza vaccination stimulates maturation of the human T follicular helper cell response

doi: 10.1038/s41590-024-01926-6

Figure Lengend Snippet: a , Gating strategy for assessing Jurkat T FH TCR cell line activation in co-culture experiments. b , Frequency of CD69 + Jurkat T cells expressing TCRs T1, T3 and T12 after co-culture with aAPCs infected with influenza PR8 for 24 hours. c , Frequency of CD69 + T1, T3 and T12 Jurkat T cell lines after co-culture with aAPCs transfected with plasmids expressing individual segments of the IAV genome compared to empty vector control for 24 hours. d , Frequency of CD69 + Jurkat T cells expressing TCRs T4, T6, T7, T9 and T10 after co-culture for 24 hours with partially HLA-matched B cells pulsed with influenza protein peptide pools. e , Frequency of CD69 + in T6 Jurkat cells stimulated as in d in the presence of antibodies blocking specific MHC class II molecules. f , Frequency of CD69 + T11 cell line co-culture with aAPCs pulsed with recombinant HA protein, PMA/ionomycin or DMSO control for 24 hours. g , PC1 scores of individual cells from the picked T FH lineages with respect to time. h , Heatmap showing the expressions of genes corresponding to the head and tail PC1 loadings in the twelve T FH clonal lineages.

Article Snippet: To identify the driving peptide motifs that triggered activation in the responding clones, a pool of overlapping oligopeptide sequences spanning M1, M2 and NP proteins of the H1N1 PR8 strain (Mimotopes) was used for the peptide mapping experiments.

Techniques: Activation Assay, Co-Culture Assay, Expressing, Infection, Transfection, Plasmid Preparation, Control, Blocking Assay, Recombinant

Journal: Cell reports

Article Title: CD62L expression marks a functionally distinct subset of memory B cells

doi: 10.1016/j.celrep.2023.113542

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Influenza A H1N1(PR8) Nucleoprotein , Sino Biological , 11675-V08B.

Techniques: Generated, Virus, Recombinant, Cell Isolation, Conjugation Assay, TA Cloning, Software

Journal: Cell reports

Article Title: CD62L expression marks a functionally distinct subset of memory B cells

doi: 10.1016/j.celrep.2023.113542

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Influenza A H1N1(PR8) Nucleoprotein (11675-V08B) was purchased from Sino Biological and conjugated using the APC Conjugation Kit-Lightning Link (Ab201807) from Abcam according to manufacturer’s instructions.

Techniques: Generated, Virus, Recombinant, Cell Isolation, Conjugation Assay, TA Cloning, Software

Journal: Virologica Sinica

Article Title: Allopregnanolone targets nucleoprotein as a novel influenza virus inhibitor

doi: 10.1016/j.virs.2023.09.003

Figure Lengend Snippet: Antiviral determination of allopregnanolone (ALLO) against influenza viruses. A The chemical structure of compound ALLO. B The dose–response effect of ALLO and oseltamivir acid upon PR8-PB2-Gluc virus replication and viabilities of MDCK cell. MDCK cell were infected with reporter influenza PR8-PB2-Gluc virus at an MOI of 0.01 in the presence of serial concentrations of ALLO. At 48 ​h post infection (h.p.i.), luciferase assay was performed to monitor virus replication. The IC 50 and CC 50 values were calculated using GraphPad Prism 5. C , D Viral titer reduction assay. MDCK cells were infected with wildtype Influenza A/PR8 (H1N1), A/Brisbane (H3N2), B/Yamagata and B/Victoria viruses at an MOI of 0.01 and treated with indicated concentrations of ALLO ( C ) and baloxivir acid ( D ) separately. At 36 ​h.p.i., the virus yields (TCID 50 /mL) were determined. Dashed lines indicate the limit of detection. Values were expressed as means ​± ​SEM (n ​= ​3). ∗∗ P ​< ​0.01; ∗∗∗ P ​< ​0.001; student's t -test. E , F In vivo antiviral evaluation. Female BALB/c mice (4–6 weeks old) were intranasally inoculated with recombinant influenza PR8-Fluc virus at a dose of 1000 TCID 50 , and received treatment with ALLO at 40 ​mg/kg/day twice daily, starting at 2 ​h before virus infection. Mice treated with vehicle only (PBS) and oseltamivir phosphate (30 ​mg/kg/day) were set as negative and positive control respectively. At day 2 p.i., bioluminescence imaging was performed ( E ) and the signal densities were analyzed ( F ). Values were expressed as means ​± ​SEM (n ​= ​3). ∗∗ P ​< ​0.01, student's t -test.

Article Snippet: To perform the thermal shift assay, 30 ​μL of solution containing 10 ​μmol/L recombinant NP protein of influenza A/PR8/Mount Sinai (Sino Biological, China) was mixed with 10 ​μL of 5× SYPRO Orange dye (MERCK, Germany) and 10 ​μL of ALLO (1.6 ​mmol/L) or DMSO as a negative control.

Techniques: Virus, Infection, Luciferase, In Vivo, Recombinant, Positive Control, Imaging

Journal: Virologica Sinica

Article Title: Allopregnanolone targets nucleoprotein as a novel influenza virus inhibitor

doi: 10.1016/j.virs.2023.09.003

Figure Lengend Snippet: Mechanistic studies for the antiviral effect of allopregnanolone (ALLO) against influenza viruses. A Schematic presentation of the time-of-addition assay. B Time-of-addition assay. MDCK cells were infected with influenza A/PR8 virus at an MOI of 0.01 and treated with ALLO or oseltamivir acid at indicated time intervals. The virus yields (TCID 50 /mL) were determined at 12 ​h post infection. C Cell-based RNA-dependent RNA polymerase (RdRp) assay. The inhibitory effect of ALLO on viral RdRp activity was tested using a mini-replicon based RdRp assay in 293T cells. Values were expressed as means ​± ​SEM (n ​= ​3). ∗ P ​< ​0.05; ∗∗ P ​< ​0.01; ∗∗∗ P ​< ​0.001; student's t -test.

Article Snippet: To perform the thermal shift assay, 30 ​μL of solution containing 10 ​μmol/L recombinant NP protein of influenza A/PR8/Mount Sinai (Sino Biological, China) was mixed with 10 ​μL of 5× SYPRO Orange dye (MERCK, Germany) and 10 ​μL of ALLO (1.6 ​mmol/L) or DMSO as a negative control.

Techniques: Infection, Virus, Activity Assay

Journal: Virologica Sinica

Article Title: Allopregnanolone targets nucleoprotein as a novel influenza virus inhibitor

doi: 10.1016/j.virs.2023.09.003

Figure Lengend Snippet: The subcellular localization of nucleoprotein (NP) during IAV infection in absence or presence of allopregnanolone (ALLO). The MDCK cells were infected with influenza A/PR8 virus and treated with (A) DMSO or (B) ALLO (40 ​μmol/L). At indicated time points, cells were fixed and stained with an anti-NP antibody (green). The nucleus was stained with DAPI (blue). The nuclear import of incoming vRNPs were indicated by red arrows, while the abnormal aggregation of newly synthesized NPs within the nucleus was indicated by yellow arrows. Scale bar ​= ​10 ​μm.

Article Snippet: To perform the thermal shift assay, 30 ​μL of solution containing 10 ​μmol/L recombinant NP protein of influenza A/PR8/Mount Sinai (Sino Biological, China) was mixed with 10 ​μL of 5× SYPRO Orange dye (MERCK, Germany) and 10 ​μL of ALLO (1.6 ​mmol/L) or DMSO as a negative control.

Techniques: Infection, Virus, Staining, Synthesized