2.4 Determination of In vitro cytotoxic effect on cultured Cells
L929/A549 cell lines were purchased from National Centre for Cell Science (NCCS), Pune, India. Cultures were maintained in Dulbecco’s modified eagles media (Himedia, India) supplemented with 10% fetal bovine serum (FBS) (Invitrogen) and grown to confluence at 37 °C and 5% CO2 in a humidified atmosphere using a CO2 incubator (NBS, Eppendorf, Germany). The cells were trypsinised (500 µL of 0.025% Trypsin in PBS/0.5 mM EDTA solution (Himedia, India) for 2 mts and passed to T flasks under aseptic condition. Test compounds were added to grown cells at a final concentration of 6.25, 12.5, 25, 50 and 100 µg/mL from a stock of 1 mg/mL and incubated for 24 hrs. The percentage difference in viability was determined by standard MTT assay (Arung et al ., 2000) after 24 hrs of incubation.