The nictitating membrane is a conjunctival fold, located at the medial canthus of the eyes, it serves to maintain healthy eyes in domestic animals. This study was conducted on 40 nictitans glands and cartilages, obtained from 20 clinically healthy local Anatolian Shepherd dogs and 20 Feral cats. The nictitans membranes were dissected, and macerated using Potassium hydroxide (KOH), and then the cartilage tissue was stained with Alcian blue stain. The samples were fixed in 10% neutral buffer formalin for histological purposes and then prepared for H&E, Verhoeff’s-Van Gieson, and Masson trichrome stains. ImageJ analyzer and AmScope^TM were used for anatomical and histological morphometrics respectively. The morphometric analysis revealed a significant difference in the cartilage morphometric dimensions between dogs and cats in correlation to their ages and weights. Both species showed a medium to strong positive correlation, but in the dogs, the cartilage body width had a low positive correlation with age and weight. The histomorphometric study of both species for the acini, interlobular septa, intralobular septa, intralobular duct, chondrocyte number, and the capsule was a significant (P>0.05) variable between species and among different groups also. We conclude that the nictitans cartilage in dogs was longer and narrower vs. to the cats, and their nictitans gland was horizontally oval-shaped while vertically oval-shaped in the cat. The nictitans gland in the dogs was composed of compound tubuloacinar serous cells that were supported by hyaline cartilage, whereas the nictitans gland in the cats was made up entirely of compound acinar units with elastic cartilage.