Colony Formation Assay: Assessing the Efficacy of Anticancer Agents on Colony-Forming Lung Cancer Cells

- The colony formation assay is an in vitro analysis of an anticancer drug on the cancer cell's growth, proliferating and differentiating into colonies in semi-solid media. To begin, seed cancer cells in the RPMI medium with growth supplement-- fetal bovine serum, or FBS, and antibiotic. Incubate the cells for the required duration.

Next, collect the desired concentration of cells in a microfuge tube and centrifuge. Re-suspend the pellet in PBS. Prepare methylcellulose-based medium, and dispense it to different tubes with FBS to promote optimal growth.

Add an appropriate amount of cell suspension in tubes, and vortex them to mix the cells with the medium. Next, add anticancer compounds in tubes, and prepare a control by adding DMSO. Vortex tubes.

Add this cell mixture to the culture plate, and incubate for the desired duration. After incubation, add methylthiazol diphenyltetrazolium bromide, or MTT, and incubate for some time. MTT reduces yellow tetrazolium salt in viable cells to purple formazan crystals. Viable colonies turn violet. Capture the images using a digital camera, and analyze the viable colonies. In the following protocol, we will study colony formation assay for asses