Biotek elisa microplate reader

To determine the effects of BTB extract on the viability of MCF-7 and MDA-MB-231 cells, an MTT assay was carried out. Cancer cells were incubated with different concentrations of extracts ranging from 25 to 500 μg/ml for 24–72 h. After the completion of the treatment time, 10 µl of 5 mg/ml MTT was added to wells and incubated for 4 h at 37 °C. Then the treatment medium was removed and 100 μl of dimethyl sulfoxide (DMSO) was added to each well to dissolve the formazan complex. The amount of colored formazan was determined by its absorbance at 570 nm in a BioTek ELISA Microplate Reader (BioTek Instrument, Inc., Winooski, VT, USA). The experiments were performed in triplicates.

T47D and TIG-1 cell lines (5 × 10³ cells/well) were seeded in 96-well plates (NEST®, NEST Scientific, USA) with RPMI and MEM complete medium, respectively, and incubated for 24 h. Cells were then treated with extracts of leaves, mesocarp, seed, and pericarp of P. macrocarpa with concentrations varying from 0 to 750 μg/mL. After 24 h of treatment, the treatment medium was removed. Then, 10 µl of WST-1 (4-[3-(4-Iodophenyl)-2-(4-nitro-phenyl)-2H-5-tetrazolio]-1,3-benzene sulfonate) reagent (Roche Diagnostics GmbH, Germany) was diluted in complete medium (1 : 10) and then added to wells and incubated for 30 min. The absorbance of the samples was measured on the BioTek ELISA Microplate Reader (BioTek Instruments, Inc., Winooski, USA) at 490 nm wavelength. The following formula determined the percentage of viability cells [12 (link)]: $\begin{array}{c}\%\text{\hspace{0.17em}cell\hspace{0.17em}viability}=\frac{\text{absorbance\hspace{0.17em}of\hspace{0.17em}treatment}-\text{absorbance\hspace{0.17em}of\hspace{0.17em}medium}}{\text{absorbance\hspace{0.17em}of\hspace{0.17em}control\hspace{0.17em}cell}-\text{absorbance\hspace{0.17em}of\hspace{0.17em}medium}}\times 100\%.\end{array}$
IC₅₀ value was calculated by a linear equation obtained from the plot between the percentages of cell viability and the log concentration of the extract. Then, the graphs were plotted in GraphPad Prism Software version 9.00 for Windows (GraphPad Software, San Diego, CA, USA).