The cells were grown at 37°C in a humidified 95%air/5%% carbon dioxide atmosphere, passaged weekly and routinely monitored for mycoplasma contamination using a detection kit (Boehringer Mannheim, Mannheim, Germany).
Cells were incubated at 37°C in 5% CO2-95% air atmosphere and passaged when near confluent monolayers were achieved using 2% trypsin-EDTA solution.
All cells were incubated at 37 °C in a 5% CO2 atmosphere and passaged once every 2 3 days.
All cells were incubated at 37°C in 5%CO2-95CO2-95%andospassagedd passaged when near confluent monolayers were achieved using trypsin-EDTA solution.
Cells were incubated at 37°C in 5% CO2-95% air atmosphere and passaged when near confluent monolayers were achieved using trypsin-EDTA solution.
Cell lines were cultured at 37°C, 5% CO2, in a humidified atmosphere and passaged for fewer than six months since receipt and stock thawing.
MDA cells obtained from the American Type Culture Collection (ATCC, USA) were cultured in RPMI-1640 medium supplemented with 10% FBS (both from Hycline) at 37°C with 5% CO2 in a humidified atmosphere and passaged every 2 3 days.
tsA-201 cells (Sigma Aldrich, St . Louis MO) were cultured in Dulbecco's Modified Eagle Medium (DMEM; Gibco-Life Technologies, Grand Island, NY) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin at 37°C in a humidified 5% CO2 atmosphere and passaged every 3 4 day.
The cells were grown in RPMI 1640 supplemented with 10%% fetal calf serum, 1 % 2 m m l-glutamine, and 1 % penicillin/streptomycin, as adherent monolayers at 310 K in a 5%% CO2 humidified atmosphere and passaged at approximately 70 80 % confluence.
All clinical S. pneumoniae strains were isolated and restreaked for single colonies on Trypticase™ Soy Agar (TSA II) supplemented with 5% sheep's blood (Becton Dickinson, Sparks, MD), and then cultured in Todd Hewitt broth (Sigma, St . Louis MO) at 37°C in a humidified 5% CO2 atmosphere for one passage followed by aliquotting and cryopreservation in 22% glycerol at –80°C.
