Culturing Vero and HeLa Cells for Infection Studies

Vero 76 cells (African green monkey kidney cells, CRL 1587 American Type Culture Collection (ATCC), Manassas, VA, USA) and HeLa cells (Homo sapiens cervix adenocarcinoma, CCL-2 ATCC) were cultured at 37°C with 5% CO₂ in growth culture medium for cell propagation. Vero growth medium consisted of Minimal Essential Medium (MEM) with Earle's salts, 25 mM HEPES, without L-Glutamine (GIBCO, Invitrogen, Carlsbad, CA, USA) supplemented with 10% fetal calf serum (FCS, BioConcept, Allschwil, Switzerland), 4 mM GlutaMAX-I (200 mM, GIBCO) and 0.2 mg/ml gentamycin (50 mg/ml, GIBCO). HeLa cell culture media were further supplemented with 1% MEM Non-Essential Amino Acids (MEM NEAA, 100x, GIBCO). Medium used for cell propagation intended for infection experiments was without gentamycin [13] (link). Cells were seeded on round glass coverslips (13 mm diameter, Sterilin Limited (Thermo Fisher Scientific), Cambridge, UK) in 24-well plates (Techno Plastic Products AG (TPP), Trasadingen, Switzerland) at a density of 3×10⁵/well in 1 mL medium for infection experiments. Infection experiments were performed when cells reached at least 90% confluency.