To describe ophthalmic examination and diagnostic values for Schirmer tear test (STT), intraocular pressure (IOP), corneal horizontal diameter (CHD), palpebral fissure length (PFL), fundoscopy, and palpebral conjunctiva microbiota from healthy giant anteaters. Twelve giant anteaters (Myrmecophaga tridactyla), 11 adults and one juvenile, five males and seven females. The animals were submitted to general anesthesia and ophthalmic evaluation with portable slit-lamp biomicroscope, Finoff transilluminator, and fundoscopy, as well as STT, bacterial culture from palpebral conjunctiva, rebound tonometry IOP, and measurement of PFL and CHD. Data compiled were analyzed with ANOVA and Tukey tests. The results (mean±standard deviation) were as follows: STT 8.04±6.21mm/min; IOP 10.92±2.45mmHg; PFL 0.75±0.11cm; CHD 0.96±0.10cm. Out of the 24 eyes swab samples for bacterial culture, 17 were positive, with three genera of Gram-positive bacteria identified Staphylococcus spp., Bacillus sp., and Corynebacterium sp. Gram-negative bacteria were not isolated from any of the samples. As conservation work in this vulnerable species continues, this report on basic ophthalmic examination and diagnostic parameters will be helpful improve their treatment and care. More ophthalmic studies are encouraged in animals within the Pilosa order.