Objective Intraocular fibrin clots caused by severe uveitis can be a sight‐threatening condition that needs to be resolved quickly and reliably. Intracameral injection of tissue‐plasminogen activator (tPA) is commonly used to resolve intraocular fibrin. However, the drug does not reach fibrinolytic concentrations after topical application. Desmoteplase (DSPA) is a structurally similar but smaller fibrinolytic agent with a higher fibrin selectivity, a longer half‐life, and better biocompatibility compared with tPA. This study was designed to evaluate the corneal and scleral permeability of DSPA in rabbits, pigs, dogs, horses, and humans ex vivo. Procedures Corneal and scleral tissues (n = 5 per group) were inserted into Franz‐type diffusion chambers and exposed to 1.4 mg/mL DSPA for 30 minutes. Drug concentrations on the receiver side were determined by liquid chromatography‐tandem mass spectrometry. Results Concentrations of DSPA after corneal and scleral permeation through fresh tissues ranged from 0.0 to 16.3 µg/mL and 0.0 to 11.4 µg/mL (rabbits), 0.3 to 5.6 µg/mL and 3.1 to 9.2 µg/mL (dogs), 2.1 to 14.9 µg/mL and 4 to 8.7 µg/mL (horses), and 0.6 to 3 µg/mL and 2.9 to 18.1 µg/mL (pigs), respectively. A concentration of 0.07‐12.9 µg/mL DSPA was detectable after diffusion through tissue culture preserved human donor bank corneas (Table 1). Conclusions Desmoteplase has the ability to permeate both cornea and sclera ex vivo in all species tested. Implications of the ex vivo permeability of DSPA suggest that in vivo permeability may be possible, and if so, it could lead to a novel topical application for lysing fibrin.