Journal: NPJ Vaccines

Article Title: Development of a Prophylactic mRNA Vaccine for Severe Fever with Thrombocytopenia Syndrome Using HSA based LNP

doi: 10.1038/s41541-026-01385-0

Figure Lengend Snippet: A Diagram showing the structure of the SFTSV Gn protein, and the modifications made to produce HSA-Gn. The HSA-Gn construct involved replacing the signal peptide (SP) with that of HSA and incorporating two sets (tmGn1 and tmGn2) of two amino acid substitutions in the transmembrane domain (TM) of the SFTSV Gn protein. B A schematic illustration of an in vitro transcription vector carrying the HSA-Gn DNA for mRNA production. C In vitro transcription of HSA-Gn using either unmodified UTP or pseudo-UTP. Linearized CLucAg containing the Cypridina luciferase gene was used as a positive transcriptional control. D Analysis of Gn and HSA-Gn proteins by Western blot. HEK293 cells were transfected with either Gn or HSA-Gn in the pSH3405 construct and cultured for 24, 48, and 72 h. Protein samples were prepared from the culture medium and cell lysate to assess the presence of Gn. Control samples (−) and (+) were derived from uninfected cells and SFTSV-infected cells, respectively. β-actin was used as an internal control. E Confocal laser scanning microscopy was used to confirm in vitro expression and the subcellular location of Gn and HSA-Gn proteins. Gn or HSA-Gn mRNA was expressed in Vero E6 cells and processed for immunofluorescence staining and analysis by confocal microscopy. Alexa Fluor 488 (Gn or HSA-Gn; green), Alexa Fluor 594 (ERGIC marker ERGIC53; red), or plasma membrane markers (PM; red) were used for staining. Following fixation, cells were stained with DAPI (blue). The cells were imaged using a Super Resolution Confocal Laser Scanning Microscope (Carl Zeiss) equipped with 405 nm, 488 nm, and 568 nm wavelengths. Shown are individual confocal sections of the red and green channels, as well as merged images including the DAPI stain. Representative images from three independent experiments are displayed. Size bar = 20 μm.

Article Snippet: The Vero E6 cell line (African green monkey kidney cells) was obtained from the Korean Cell Line Bank and maintained in Dulbecco’s Modified Eagle Medium (DMEM; GIBCO) supplemented with 10% heat-inactivated fetal bovine serum (FBS; GIBCO) and 1% penicillin–streptomycin (GIBCO) at 37 °C in a humidified atmosphere with 5% CO2 until 80% confluence was reached.

Techniques: Construct, In Vitro, Plasmid Preparation, Luciferase, Control, Western Blot, Transfection, Cell Culture, Derivative Assay, Infection, Confocal Laser Scanning Microscopy, Expressing, Immunofluorescence, Staining, Confocal Microscopy, Marker, Clinical Proteomics, Membrane, Laser-Scanning Microscopy