3.5.5. Cell Viability by Sulphorhodamine B (SRB) Test

For the analysis of SRB, the method described by Papazisis et al. [59] was used. In brief, Calu-3 cell line was treated with each sample for 72 h and the supernatant was then discarded. The cells were washed with 200 μL of sodium phosphate buffer at pH = 7.4. Then, 200 μL of 10% cold trichloroacetic acid was added and the plates were placed in the refrigerator for 30 min to fix the cells. Subsequently, the cells were washed three times with 200 μL of distilled water and maintained for 24 h at room temperature (20 ± 2 ◦C) to dryness. Later, 200 μL of 0.2% SRB solution was added and kept for 30 min. The plates were then washed five times with 200 μL of 1% acetic acid and again dried for 30 min. Finally, 150 μL of 10 mmol.L−<sup>1</sup> TrisBase was added and the spectrophotometric absorbance reading was performed at a wavelength of 432 nm in a microplate reader (μQuant, BioTek). Equation (2) was also used for calculating cell viability (%).
