Objective To validate the gross tumor volume (GTV) delineation in head and neck cancer based on magnetic resonance imaging (MRI) or computed tomography (CT) by cross-sectional autopsy, and to provide a basis for clinical evaluation of the two imaging methods. Methods Ten New Zealand rabbits were selected for transplantation of VX₂ carcinoma cells, and a head and neck cancer model was successfully established in six rabbits. Each rabbit was fixed and received MRI scan and CT scan in the same body position. Then, they were sacrificed and fixed in gelatin solution (－70℃) for 72 h;all cryopreserved rabbits underwent cross-sectional autopsy using a jig saw, with the same position and sectional thickness as in MRI scan and CT scan, and cross-sectional autopsy images were obtained using a digital single-lens reflex camera. GTVs were separately delineated based on CT, MRI, and cross-sectional autopsy images. The GTV_MRI, GTV_CT, GTV_SA, and volume difference ratios (VDRs) were calculated;two-way classification ANOVA and paired t-test were used for difference analyses. Results The mean values of GTV_MRI, GTV_CT, and GTV_SA were 8.20±2.56, 8.40±2.20, and 8.11±2.88 cm³, respectively, without significant differences among them (F=0.06,P=0.943). The mean values of VDR_MRI-SA and VDR_CT-SA were 0.180±0.060 and 0.309±0.091, respectively, with a significant difference between them (t=7.49, P=0.001). Conclusion The GTV delineation based on MRI is more accurate than that based on CT in head and neck cancer.