To conduct a HET-CAM assay, fertilized chicken eggs are incubated until the chorioallantoic membrane is well-developed. A small window is made in the eggshell to expose the membrane, and the test substance is applied. Observations are made over a specific timeframe to record any irritation responses such as hemorrhage, coagulation, or lysis. These responses are then scored to determine the irritation potential of the substance.
