cchfv gc protein human fc tag  (Native Antigen Inc)

Journal: Pathogens

Article Title: Enhanced Seroconversion to West Nile Virus Proteins in Mice by West Nile Kunjin Replicon Virus-like Particles Expressing Glycoproteins from Crimean–Congo Hemorrhagic Fever Virus

doi: 10.3390/pathogens11020233

Figure Lengend Snippet: Expression of Crimean–Congo hemorrhagic fever virus (CCHFV) glycoproteins Gn and GC proteins using the West Nile Kunjin (WNV KUN ) replicon. ( A ) The reporter luciferase (Luc) gene was substituted with genes encoding CCHFV Gn and Gc in the WNV KUN DNA replicon. In short, the replicon is driven by the cytomegalovirus (CMV) promoter expressing an open reading frame flanked by the 5′- untranslated region (UTR) and the 3′-UTR comprising: first, partial capsid (C) gene fused in frame with the Luc, the foot and mouth disease virus autoprotease 2a (FMDV2A) then partial envelop (E) gene, and all the nonstructural proteins. The hepatitis delta virus antigenomic ribozyme (HDVr) sequence was inserted immediately downstream of the WNV KUN 3′-UTR followed by the Simian virus 40 (SV40) polyadenylation signal (pA). ( B ) Immunoblotting of cell lysates 2 days after transfection of the WNV KUN CCHFV Gn–Gc replicons versus the WNV KUN Luc replicon into the BHK-21 cell line expressing the WNV KUN C–prM–E.

Article Snippet: The following proteins and antibodies were used in this study: the WNV NY99 E protein, the WNV NY99 NS1 protein, the CCHFV Gn protein, the CCHFV Gc protein human Fc tag, the mouse anti-CCHFV Gn (The Native Antigen Company, Oxford, UK), the mouse anti-flavivirus NS1 (Abcam, Cambridge, UK), the mouse J2 anti dsRNA (Scicons, Szirák, Hungary), the mouse anti-CCHFV Gn–Gc (Friedrich-Loeffler-Institute, Greifswald, Germany), the Alexa Fluor 594-conjugated anti-mouse goat antibody (Invitrogen, Vilnius, Lithuania), and the HPR-conjugated anti-mouse goat antibody (Invitrogen).

Techniques: Expressing, Virus, Luciferase, Sequencing, Western Blot, Transfection

Journal: Pathogens

Article Title: Enhanced Seroconversion to West Nile Virus Proteins in Mice by West Nile Kunjin Replicon Virus-like Particles Expressing Glycoproteins from Crimean–Congo Hemorrhagic Fever Virus

doi: 10.3390/pathogens11020233

Figure Lengend Snippet: Immunofluorescence labeling of BHK-21 C-prM-E cells after transfection with the Gn–Gc replicon, followed by two consecutive cycles of RVP infection into naïve BHK-21 cells. The cells were visualized with the antibodies anti-CCHFV Gn–Gc (red), anti-WNV KUN NS1 (red), and anti-dsRNA (red). The nucleus was counterstained with DAPI (blue). Bar scales represent 20 µm.

Article Snippet: The following proteins and antibodies were used in this study: the WNV NY99 E protein, the WNV NY99 NS1 protein, the CCHFV Gn protein, the CCHFV Gc protein human Fc tag, the mouse anti-CCHFV Gn (The Native Antigen Company, Oxford, UK), the mouse anti-flavivirus NS1 (Abcam, Cambridge, UK), the mouse J2 anti dsRNA (Scicons, Szirák, Hungary), the mouse anti-CCHFV Gn–Gc (Friedrich-Loeffler-Institute, Greifswald, Germany), the Alexa Fluor 594-conjugated anti-mouse goat antibody (Invitrogen, Vilnius, Lithuania), and the HPR-conjugated anti-mouse goat antibody (Invitrogen).

Techniques: Immunofluorescence, Labeling, Transfection, Infection

Journal: Pathogens

Article Title: Enhanced Seroconversion to West Nile Virus Proteins in Mice by West Nile Kunjin Replicon Virus-like Particles Expressing Glycoproteins from Crimean–Congo Hemorrhagic Fever Virus

doi: 10.3390/pathogens11020233

Figure Lengend Snippet: CCHFV Gn–Gc RVPs induced seroconversion to CCHFV Gn and Gc and enhanced seroconversion to WNV NY99 NS1, E. ( A ) Schematic illustration of the mice immunization schedule. Mice were subcutaneously immunized three times at weeks 0, 2, and 5 with RVPs expressing CCHFV Gn–Gc (6 mice), Luc (6 mice), or phosphate-buffered saline (PBS) (3 mice). ( B ) Mouse weight from one week before the experiment to the mouse-euthanized day. Mice sera from the study groups were diluted and assayed with enzyme-linked immunosorbent assays (ELISA) to measure antibody titers against CCHFV Gn ( C ), CCHFV Gc ( D ) WNV NY99 E ( E ), and WNV NY99 NS1 ( F ). The end-point titers were determined as there was no difference in the measured optical density values at 450 nm (OD450) between the vaccinated group and the control group. The experiments were conducted with two technical repeats. The p values are indicated using * p < 0.05 and ** p < 0.01. ( G ) Serum titers that elicited 50% reduction in the WNV KUN plaque number. Sera from experimented animal were combined before assaying.

Article Snippet: The following proteins and antibodies were used in this study: the WNV NY99 E protein, the WNV NY99 NS1 protein, the CCHFV Gn protein, the CCHFV Gc protein human Fc tag, the mouse anti-CCHFV Gn (The Native Antigen Company, Oxford, UK), the mouse anti-flavivirus NS1 (Abcam, Cambridge, UK), the mouse J2 anti dsRNA (Scicons, Szirák, Hungary), the mouse anti-CCHFV Gn–Gc (Friedrich-Loeffler-Institute, Greifswald, Germany), the Alexa Fluor 594-conjugated anti-mouse goat antibody (Invitrogen, Vilnius, Lithuania), and the HPR-conjugated anti-mouse goat antibody (Invitrogen).

Techniques: Expressing, Saline, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Side-by-side evaluation of two mouse models for Crimean-Congo Hemorrhagic Fever Virus infection

doi: 10.1101/2025.08.20.671421

Figure Lengend Snippet: IFNAR -/- mice were infected with either 100 TCID 50 CCHFV Afg09 (purple, n = 8) or Hoti (green, n = 6). The clinical score (A) , summarizing appearance, behaviour, body weight (B) , and body temperature (C) , was monitored daily. After reaching the clinical endpoint (score of ≥10 or ≥6 on two consecutive days), the mice were sacrificed or found dead (grey). Dotted lines mark the clinical endpoints.

Article Snippet: Microtiter plates were coated with CCHFV glycoprotein (Gc) protein (The Native Antigen Company, REC31696) diluted to 0.5 μg/ml in phosphate buffered saline (PBS) and incubated for 20 hours at 4°C.

Techniques: Infection

Journal: bioRxiv

Article Title: Side-by-side evaluation of two mouse models for Crimean-Congo Hemorrhagic Fever Virus infection

doi: 10.1101/2025.08.20.671421

Figure Lengend Snippet: IFNAR -/- mice were infected with either 100 TCID 50 CCHFV Afg09 (purple, n = 8) or Hoti (green, n = 6). A: CCHFV genome copies were measured by RT-qPCR in organs and serum samples (day 3 and final) of individual mice. B: Infectious CCHFV were determined by TCID 50 assay in organs and final serum samples of individual mice. Each data point represents a sample from an individual animal, data are shown as the means ± SD. Datasets were analyzed using the Šídák’s multiple comparisons test (A) or Tukey’s multiple comparisons test (B). Asterisks indicate statistical significance as detailed between CCHFV Afg09 and Kosovo Hoti group: ∗p ≤ 0.05; ∗∗∗∗p < 0.0001.

Article Snippet: Microtiter plates were coated with CCHFV glycoprotein (Gc) protein (The Native Antigen Company, REC31696) diluted to 0.5 μg/ml in phosphate buffered saline (PBS) and incubated for 20 hours at 4°C.

Techniques: Infection, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Side-by-side evaluation of two mouse models for Crimean-Congo Hemorrhagic Fever Virus infection

doi: 10.1101/2025.08.20.671421

Figure Lengend Snippet: IFNAR -/- mice were infected with either 100 TCID 50 CCHFV Afg09 (purple) or Hoti (green). A: Final sera (Afg09 n = 6, Hoti n = 6) were analyzed with two virus-specific ELISAs using either inactivated CCHFV, measuring NP-specific antibodies or the major glycoprotein Gc for coating. Monoclonal antibodies detecting Gc or N protein were used as controls (not shown). Dotted lines indicate the respective lower limit of detection. B: Liver enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were analyzed in final serum samples (Afg09 n = 5, Hoti n = 3). Dotted lines indicate the physiological range of healthy C57BL/6 mice.

Article Snippet: Microtiter plates were coated with CCHFV glycoprotein (Gc) protein (The Native Antigen Company, REC31696) diluted to 0.5 μg/ml in phosphate buffered saline (PBS) and incubated for 20 hours at 4°C.

Techniques: Infection, Virus, Bioprocessing

Journal: bioRxiv

Article Title: Side-by-side evaluation of two mouse models for Crimean-Congo Hemorrhagic Fever Virus infection

doi: 10.1101/2025.08.20.671421

Figure Lengend Snippet: Liver, spleen and CNS of CCHFV Afg09 (purple) or Hoti-infected (green) IFNAR -/- mice were pathologically examined post-mortem. A: Shown are exemplary images of organ histopathology, H&E: hematoxylin and eosin staining. ISH: In situ Hybridization. Arrows indicate multifocal liver necrosis with loss of tissue architecture and lymphohistiocytic infiltrates. Arrowheads indicate apoptosis in germinal follicle centers. Scale bars: 100 µm. B: Quantification of ISH indicating percentage area of CCHFV-positive organ sections. Each data point represents a sample from an individual animal, horizonal lines represent the means ± SD. Datasets were analyzed using the Šídák’s multiple comparisons test. Asterisks indicate statistical significance as detailed between CCHFV Afg09 and Kosovo Hoti group: ∗∗∗∗p < 0.0001; ns = not significant.

Article Snippet: Microtiter plates were coated with CCHFV glycoprotein (Gc) protein (The Native Antigen Company, REC31696) diluted to 0.5 μg/ml in phosphate buffered saline (PBS) and incubated for 20 hours at 4°C.

Techniques: Infection, Histopathology, Staining, In Situ Hybridization