Chicken Chorioallantoic Membrane-based Cancer Modeling: An In Ovo Model to Study Cancer Cell Tumorigenesis and Metastasis

To prepare a cancer cell suspension for transplantation, harvest the cells from the cancer cell culture of interest and sediment the cells by centrifugation. Aspirate the supernatant and mechanically resuspend the cells in the residual medium. Place the cell suspension on ice and use a pipette to measure the volume of cells in medium. Then, resuspend the cells at a concentration of 1 to 2 million cells in 20 to 100 microliters of medium, supplemented with 2.7 to 4 milligrams per milliliter extracellular matrix and any growth factors or other additives of interest.

For cancer cell implantation, using a non-stick ring, place up to 6 eggs to be implanted onto the egg rack and roll the edges of the transparent film dressing toward the open window to remove the film from the shells. Check whether the eggs are viable and healthy. Ideal eggs will have a large vessel with smaller branching vessels within the center of the opened area.

Using curved iris forceps, place a sterile non-stick ring onto the CAM over the vessel, ideally over a branch point. Use a sterile glass stir rod to gently abrade the CAM and pipet the cancer cell suspension into the center of the ring. When all of the cells have been delivered, seal the opening with a one quarter piece of 6 by 7 centimeter transparent film dressing and label the egg with an appropriate implant designation. Then, place the egg securely in the incubator, with the opening of the shell facing upright.