JoVE Encyclopedia of Experiments
Cancer Research
0 views • 3:57 min • April 30th, 2023
Sulforhodamine B or SRB assay allows cell density determination based on the measurement of cellular protein content. To begin the assay, pipette a suspension of desired drug-sensitive parental cancer cells and their drug-resistant variant into wells of a multi-well plate.
Incubate to allow the cells to adhere to the plate. Treat both the cell types with a range of increasing concentrations of the specific drug and incubate.
In culture, the drug-sensitive cancer cells are susceptible to the drug molecules and undergo cell death even at low concentrations. In contrast, the drug-resistant cancer cells withstand the effect of drug molecules and survive and proliferate even at higher drug concentrations.
Next, add an appropriate concentration of trichloroacetic acid or TCA to the culture. TCA - a mild acid - permeabilizes the cells and disrupts the electrostatic interactions and native protein conformations, leading to protein denaturation.
Incubate with Sulforhodamine B solution. Under mild acidic conditions, sulforhodamine B - a protein-binding dye - binds stoichiometrically to basic amino acid residues of proteins.
Rinse with acetic acid to remove any unbound dye. Now, treat the sa
View the full transcript and gain access to thousands of scientific videos