Chick Chorioallantoic Membrane or CAM Tumor Model: An In Ovo Method to Simulate Perineural Invasion of Tumor Cells

Abstract

Source: Schmidt, L. B. et al. The Chick Chorioallantoic Membrane In Vivo Model to Assess Perineural Invasion in Head and Neck Cancer. J. Vis. Exp. (2019)

In this video, we describe the method to co-culture dorsal root ganglia and cancer cells on chick chorioallantoic membrane or CAM to study perineural invasion.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Grafting DRG on the CAM (estimated timing: 40 min, day 8) Prepare a 6 cm culture dish with HBSS medium to wash the DRGs before implantation. Bring the prepared eggs to the cell culture laminar flow cabinet. Remove the paraffin membrane from the egg (Figure 1A). With fine sterile forceps, …

Representative Results

Figure 1: Grafting of DRG, cells, and harvesting of CAM: On day 8: A. CAM easily observed after paraffin wax membrane removal. B-C. With fine forceps, DRG is placed onto the CAM. D. Egg is covered with film dressing and put in the incubator; arrows point to the openings that are covered. On day 10: E-F….

Materials

Egg incubator	GQF Digital Sportsman	#1502	Egg incubator equipped with automatic rotator digital thermostat temperature and humidity controls
Engraving cutter	DREMEL	#108	Used to drill the egg shell
Extra fine Graefe forceps, curved	Fine Science Tools (FST)	#11151-10	Used to graft DRG on to the CAM on day 8 and to harvest CAMtissue on day 17
Extra fine Graefe forceps, straight	Fine Science Tools (FST)	#11150-10	Used to graft DRG on to the CAM on day 8 and to harvest CAM tissue on day 17
Fertilized Lohmann White Leghorn eggs			Fertilized eggs at early fertilization days preferably on first day postfertilization. Eggs used in this protocol are from Michigan State University Poultry Farm.
Filter Forceps	EMD Millipore	#XX6200006P	Blunt forceps used to remove the egg shell
HBSS (1x)	Gibco	#14025-092	Hank`s Balanced Salt Solution
Paraffin wax membrane	Parafilm laboratory film	#PM-996	Used to temporarily cover the egg openings until DRG grafting on day8
PBS (1x) pH 7.4	Gibco	#10010-023	Phosphate Buffered Saline
Tegaderm Transparent Film Dressing	3M	#9505W	Sterile, 6x7cm used to cover the egg openings during incubation