3D-cocultuur met directe interactie: kankercellen co-cultiveren met monocyten om hun interactie te bestuderen

To begin, add DQ collagen IV, a fluorescent dye quenched protein substrate into an extracellular matrix extract or ECME. Pour this extract into each well of a two well slide. Label the first well as co-culture, second as control. Add an appropriate amount of fluorescently labeled cancer cell suspension into wells, and incubate the slide at 37 degrees Celsius for the desired time to promote cell attachment.

Add a required amount of monocytes cell suspension to the first well. Let the ECME solidify at 37 degrees Celsius. Pour equal ratios of cancer cell and monocyte culture media into each well and incubate the slide at 37 degrees Celsius for the desired duration in a carbon dioxide environment. In the co-culture well, cancer cells secrete proteins such as monocytes chemoattractant protein to attract monocytes.

In contrast, monocytes produce matrix metalloproteinase or MMP, to degrade the collagen, promoting cancer cells invasion. Observe the slide under a confocal microscope. The DQ collagen degradation emits fluorescence which can be seen more in the co-culture cells indicating cell interaction, while less in control cells. In the following protocol, we will co-culture breast cance