Dulbecco s modified

HeLa cells (Sigma-Aldrich) were maintained in Dulbecco’s modified Eagle’s medium, supplemented with 10% fetal bovine serum (Biowest SAS, France), 4 mM L-glutamine, 100 U/ml penicillin and 100 µg/ml streptomycin solution. Cells were cultured in a humidified atmosphere containing 5% CO₂ at 37 °C. On reaching 80% confluence, cells were detached every 3–4 days using 0.05% (w/v) trypsin, 0.02% (w/v) EDTA solution.

Primary cultures of neonatal rat cardiomyocytes were prepared as described as previously [35 (link)]. In brief, neonatal ventricular myocytes from 1- to 2-day-old Sprague-Dawley rats were sacrificed by decapitation and hearts were immediately removed and subjected to Percoll gradient centrifugation and differential plating to enrich the cardiac myocyte population and to deplete non-myocytes. Cardiomyocytes were cultured in a mixture of Dulbecco’s modified Eagle’s medium (DMEM) and M199 with 10% fetal bovine serum (FBS; Biowest, Nuaille, France).

HeLa cells
(ECACC 93021013 human, cervix, epitheloid, and carcinoma) were maintained
in a humidified incubator (37 °C, 5% CO₂) in Dulbecco’s
modified Eagle’s medium, supplemented with 4 mM L-glutamine,
10% fetal bovine serum (Biowest SAS, France), 100 U/mL penicillin,
100 μg/mL streptomycin solution, and 0.25 μg/mL amphotericin
B. Cell adhesion assay buffer was prepared by adding 20 mM 4-(2-hydroxyethyl)-1-piperazine
ethanesulfonic acid (HEPES, Sigma-Aldrich) to Hank’s balanced
salt solution (HBSS, Sigma-Aldrich), pH 7.0.^{1 (link)}

Virus infections were performed on Vero ATCC CCL-81 or SH-SY5Y ATCC CRL-2266 cells (ATCC, Manassas, VA). Vero cells were cultured in minimal essential medium (MEM) (Corning, Manassas, VA) supplemented with 10% fetal bovine serum (Gibco, Life Technologies, Paisley, UK), and SH-SY5Y cells were grown in Dulbecco’s modified Eagle’s medium (DMEM)–F-12 medium (Biowest, Nuaillé, France) supplemented with 10% fetal bovine serum. A penicillin-streptomycin mixture (Corning) and l-glutamine (Corning) were also added to cell cultures. WNV New York 99 (60 (link)), USUV SAAR 1776 (60 (link)), American ZIKV PA259459 (61 (link)), a cell-passaged derivative of DENV-2 16681 (16 (link)), VSV Indiana (61 (link)), and CVB5 Faulkner (62 (link)) were used. Procedures for infections in liquid medium and virus titrations in Vero cells in semisolid agar medium were previously described (62 (link), 63 (link)). WNV, USUV, ZIKV, VSV, and CVB5 titers were determined at 24 h postinfection (p.i.). DENV-2 titers were determined at 48 h p.i. A multiplicity of infection (MOI) of 1 PFU/cell was used for all experiments unless otherwise indicated.