JoVE Encyclopedia of Experiments
Cancer Research
0 views • 3:31 min • April 30th, 2023
- Sometimes, a stem or progenitor cell undergoes mutations, giving rise to a leukemic stem cell. In these cells, certain proteins are upregulated, and using anticancer compounds against such proteins can help control cells' growth. The effect of these anticancer compounds can be analyzed using CFU assay by monitoring the number and size of colonies formed.
Begin by taking progenitor cells from a leukemic mouse and diluting them in a suitable culture media. Add this cell suspension along with the appropriate anticancer compound to methylcellulose containing mouse-specific cytokines, and pour it into a Petri dish. Incubate this mixture till the colonies appear. Methylcellulose is a chemically inert, semi-solid matrix that prevents cell migration while cytokines stimulate progenitor cells to differentiate and form colonies containing recognizable progeny.
Next, stain the colonies using iodonitrotetrazolium chloride, or INT, and incubate overnight till the colonies turn dark reddish brown. Mitochondrial enzymes reduce yellow-colored INT to reddish brown-colored formazan dye. Finally, identify and count the colonies present on the plate. In the following protocol, we will perform a CFU
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