Assessment of In Vitro Cytotoxic Activity of Fungal Extracts by MTT Assay

The cytotoxicity of the extracts was examined by the MTT assay by determining the viability of cells (Kuriakose et al., 2014). Briefly, HeLa cells were grown and maintained in DMEM medium and supplemented with 10% fetal bovine serum, 100 U/ml penicillin, 100 mg/ml streptomycin, and 1% (v/v) GlutaMAX and incubated at 37°C in a 5% CO₂ of the humidified chamber. 1 × 10⁴ cells/ml HeLa cells in 100 µL of DMEM were seeded in 96-well plates and incubated for 24 h. Thereafter, the cells were treated with fungal crude extracts at a range of concentrations (10, 50,100, and 200 μg/ml) in 100 µL media and incubated for 48 h, respectively. At the end of 48 h, 10 µL of MTT reagent (5 mg/ml) was added to each well. After 2 h of incubation at 37°C, the plates were emptied and MTT reagent was removed. To the each well, dimethyl sulfoxide (100 µL) was added and then the absorbance was measured at 595 nm wavelength with an ELISA reader Infinite M2000 Pro™ (Tecan, Crailsheim, Germany) (Kumari et al., 2018). The IC₅₀ values were calculated for each fungal extract. All the assays were performed in triplicates and results were presented as mean ± SD. Aspergillus sp. extract demonstrated the highest cytotoxicity and was, hence, chosen for further investigation.