Vero e6 cells

HAZV Strain JC 280 as well as SW13 cells were kindly provided by Ali Mirazimi, Karolinska Institutet, Stockholm, Sweden. For indirect immunofluorescence assays, Vero E6 cells (Collection of Cell Lines in Veterinary Medicine, Friedrich-Loeffler-Institut, Greifswald, Germany) were employed. SW13 cells were cultivated in L-15 Medium (Sigma Aldrich, Darmstadt, Germany) supplemented with 5% fetal calf serum (FCS), whereas Vero E6 cells were cultivated in Modified Eagles Medium (MEM, Collection of Cell Lines in Veterinary Medicine, Friedrich-Loeffler-Institut, Greifswald, Germany) supplemented with 10% FCS. Virus stocks were grown in SW13 cells. Growth kinetics were performed revealing maximum viral titers 48 h post-infection (up to 10^7.8 TCID₅₀/mL). A cytopathic effect becomes first visible 3 days post-infection (dpi).

Acrylamide (AAm) (purity > 98%), 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPSA) (purity > 99%), methacrylic acid (MA) (purity > 99%), ammonium persulfate (purity > 98%), N,N,N’,N’-tetramethylethylenediamine (purity > 99%), 1-vinyl-2-pyrrolidone (VP) (purity > 99%), and CPC (purity > 98%, critical micelle concentration (CMC) in water 0.9 mM [42 (link),43 (link),44 (link)]) were provided by Sigma Aldrich (St. Louis, MO, USA). N,N′-methylenebisAcrylamide (purity > 99%), azobisisobutyronitrile (purity > 98%), and polyethylene glycol (PEG) with a molar mass of 1000 g/mol were obtained from Fluka (New York, NY, USA). Isopropanol was purchased from Acros Organics (Geel, Belgium). All chemicals were used as received. The solutions were prepared using distilled deionized water obtained by MilliQ system (Millipore, Burlington, MA, USA).
Vero E6 cells (ATCC, Manassas, VA, USA; catalog number CRL-1586) were cultured in high glucose Dulbecco’s modified Eagle’s medium (DMEM; Sigma-Aldrich, St. Louis, MO, USA) supplemented with 5% fetal calf serum (FCS), 2 mM L-glutamine, and antibiotics (150 u/mL penicillin and 150 u/mL streptomycin) at 37 °C in 5% CO₂.
The stock of SARS-CoV-2 (strain HCoV-19/Russia/Moscow-PMVL-12/2020 (EPI_ISL_572398) isolated from a patient) was the culture liquid withdrawn from cultures of the infected Vero E6 cells.

Vero-E6 cells (ATCC-CRL-1586) were maintained in Dulbecco’s Modified Eagle Medium (Sigma, Munich, Germany) supplemented with 10% fetal calf serum and 1% penicillin–streptomycin and cultured at 37 °C in a humidified incubator with 5% CO₂. SARS-CoV-2, isolated from the first Swedish patient, was received from the Public Health Agency of Sweden.

African green monkey kidney epithelial Vero E6 cells were purchased from the American Type Culture Collection (ATCC CRL-1586; Manassas, VA, USA), and the SARS-CoV-2, KCDC03 (isolated from Korean COVID-19 patient in 2020 and belonging to the A lineage of early Chinese strains), was kindly provided by the National Culture Collection for Pathogens in Korea. The Vero E6 cells were grown in Dulbecco’s modified Eagle’s medium (DMEM, Gibco) supplemented with 5% FBS (Gibco) and 1% antibiotic-antimycotic (Gibco, Cat No. 15240-062) at 37 °C with 5% CO₂. The SARS-CoV-2 KCDC03 strain was propagated in Vero E6 cells in the presence of 1 μg/ml tosyl phenylalanyl chloromethyl ketone (TPCK) trypsin (Sigma-Aldrich, Cat No. 4370185).

The Bangladeshi strain of the Nipah virus (NIV) was obtained from the Viral Special Pathogens Branch of the CDC, was originally isolated from a fatal human infection in Bangladesh in 2004, and was passaged in Vero E6 cells a total of three times [16 (link)]. Cedar virus (CedPV) was obtained from CSIRO (Geelong, Australia) and was isolated from bat urine inoculated onto primary bat kidney cells, followed by passaging on Vero E6 cells (ATCC) [7 (link)]. Both viruses were propagated on Vero E6 cells at RML in Dulbecco’s minimal essential medium (DMEM) (Sigma) supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 50 IU/mL penicillin, and 50 µg/mL streptomycin (Life Technologies, Carlsbad, CA, USA).
BHK-21 cells (CCL-10, ATCC, Manassas VA, USA) were propagated in 24-well tissue culture plates and inoculated with CedPV or NiV (or mock) with a MOI of 1.0 or 0.1 for 1 h, washed twice in DMEM, and incubated at 37 °C, 5% CO₂ for the indicated amount of time.
Human lung blood microvascular endothelial cells (LBMVEC) (CC-2527, Lonza, Walkersville MD, USA) were maintained in an EGM-2 medium (Lonza). The cells were seeded in 24-well plates one day prior to inoculation with CedPV or NiV with a MOI of 0.1. At the indicated time points, supernatants were collected for virus quantitation and the cells were lysed in buffer RLT (Qiagen) to quantitate viral or cellular RNA.