C myc tag

The cells were cultured on glass coverslips (Millipore, Billerica, MA, USA) until they were semi-confluence and then incubated with HTNV for 60 min (moi = 1). At the indicated times post-HTNV infection, the cells were fixed with 4% PFA, incubated with 0.3% Triton X-100 (Sigma-Aldrich, St. Louis, MO, USA), and blocked with 5% BSA for 1 h. Following incubation with a mouse monoclonal antibody against c-myc-tag (Sigma-Aldrich, St. Louis, MO, USA, Sigma-Aldrich Cat# M5546), IFITM3, lysosome-associated membrane glycoprotein 1 (LAMP1, Cell Signaling Technology, Danvers, MA, USA), or HTNV NP at 37°C for 2 h, the cells were washed and incubated with anti-rabbit Ig conjugated to Alexa 555 and anti-mouse Ig conjugated to Alexa 488 (Abcam, Cambridge, MA, USA) secondary antibodies at room temperature for 1 h. The nuclei were counterstained with DAPI. An Olympus BX51 fluorescence microscope system and FV1000 confocal microscopy system (Olympus, Tokyo, Japan) were used to capture the images.

The protein concentration of purified VLPs was determined by Bradford assay (Bio-Rad), and the BCA assay (Thermo Fisher) was used for samples of the serum purification process. For verification of sample quality and purity, EBOV VLP samples containing 10 µg total protein or NiV VLP samples containing 3 µg total protein were separated by 10% reducing SDS-PAGE and antiserum samples containing 10 µg total protein were separated by 8% non-reducing SDS-PAGE and stained with Coomassie blue (Fisher Bioreagents) or transferred to polyvinylidene difluoride (PVDF) membranes (Millipore) for Western blot analysis. Blots were stained with a polyclonal goat antiserum against EBOV virus^{72 (link)} at a working solution of 1:1000 or monoclonal antibodies against FLAG-tag, HA-tag, or c-myc-tag (all Sigma) at the recommended dilution to detect the respective epitope-tagged NiV proteins.