Soft Agar Colony Formation Assay: een methode om effecten van nieuwe verbindingen op de proliferatie van kankercellen te testen

To begin, take an appropriate concentration of molten agarose in a conical tube and add a desired amount of warm culture medium to it. Gently invert to mix the contents. Add this mixture into each well of a six-well plate, and allow it to solidify. This is the bottom layer. Next, treat cancer cells with the compound of interest and add the required concentration of agarose.

Now, pour this mixture containing the desired number of cells per milliliter into each well. This is a cell-containing layer. Once the medium solidifies, incubate the plate at 37 degrees Celsius for a week. Prepare a feeder layer by mixing agarose to the medium and supplement this mixture with the compound of interest. Gently add this feeder layer into each well, allow it to solidify, and incubate the plate at 37 degrees Celsius.

Repeat this feeding procedure weekly to replenish the cells with the medium until colonies form. If the compound of interest is an inhibitor of tumorigenicity, it will suppress cells growth resulting in few colonies in the wells. In the following protocol, we will perform a soft agar colony formation assay to study the effect of PADI inhibitors on the tumorigenicity of breast cancer cel