Write a procedure that could be used to generate these resul

Write a procedure that could be used to generate these results using a MTT assay, and explain your interpretation of this experiment (2 points). (Note: Stock Dox concentration is 100 micromolar)

1201 100 80 60 40 20 Cytotoxicity of Doxorubicin on KB cells D2 days O3 days e4 days RPM1 Dox 250 Dox 300 Dox 150 Treatments (nM)

Solution

Answer : Cytotoxicity of Doxorubicin on KB cells

Doxorubicin (adriamycin) is a potent cytotoxic and DNA-damaging anthracycline chemotherapeutic agent The present data investigating the evaluation of cytotoxicity of Doxorubicin on KB cells.

Procedure:

The KB cells (reached 90% confluence in T25 flasks) were trypsinized. After washing with PBS, it has to be counted in a Neubauer slide using the trypan-blue dye-exclusion test. Then seed the cells into 96-well plates at a density of 5×10 ³ cells/well for 48 h. Different concentrations of 150, 250 and 300 nM doxorubicin were treated with KB cells. Take RPMI culture medium as a control. Determine the growth rate by MTT assay after incubation for two, three, and four days.

MTT assay: Each well was incubated with 4 mg/ml of MTT in PBS at 37°C in the CO ₂ incubator. MTT were discarded from the wells after 3 h and 100 l of DMSO were added to solubilize the formazan crystals. Absorbance was recorded at a test wavelength of 540 nm. Percent survival was calculated.

Result:

The results show the antiproliferative effect of doxorubicin on KB cells was time- and dose-dependent. The percentage of cell survival decreases as the time and dose increases.