Chick Chorioallantoic Membrane Harvest: A Method to Isolate Tumor Graft-bearing Chorioallantoic Membrane from Chicken Egg

The chorioallantoic membrane or CAM is an extraembryonic membrane, adhering closely to the inner shell membrane of an intact egg. CAM is an ideal system for tumor implantation and modeling because it is a naturally immunodeficient and highly vascularized membrane.

To harvest a tumor-bearing CAM, first, take a viable fertilized chicken egg having a fully developed CAM previously transplanted with a tumor. Add paraformaldehyde to the CAM. Paraformaldehyde crosslinks the membrane proteins, making the membrane stiffer to facilitate its harvesting.

Next, cut around the egg to separate the upper portion of the eggshell from the remainder of the egg, with the tumor-bearing CAM still attached to the removed shell. Hold and lift the CAM at one edge to smoothly detach it from the eggshell.

Thereafter, place the harvested CAM in paraformaldehyde with the tumor graft facing upwards. Paraformaldehyde preserves the morphology of the CAM and tumor graft, thereby preparing the harvested membrane for various downstream analyses.

At the end of the incubation, place the eggs onto the laboratory benchtop. Use a syringe to perforate the film dressing of each egg and apply about 300 microliters of 4% paraformaldehyde over each chorioallantoic membrane to slightly stiffen the tissue to facilitate the harvesting process.

Using scissors, remove the upper half of the eggshell with the chorioallantoic membrane attached. Reduce the size of the shell to approximately 3 centimeters in diameter, keeping the portion of the chorioallantoic membrane where dorsal root ganglion and tumor cells were grafted within the center of the shell fragment. Then, grasp the chorioallantoic membrane with fine forceps and detach the tissue from the eggshell for transfer into one well of a 6-well plate containing 2 milliliters of 4% paraformaldehyde per well.