A 32-year-old lactating woman with open-angle glaucoma used timolol maleate 0.5% eye drops twice daily to her right eye for 6 months. Four milk samples were collected over a span of 6 days. Timolol maleate milk levels were examined by liquid chromatography tandem mass spectrometry and found to be at a mean of 0.12 ng/mL (range, 0 to 0.37 ng/mL). At this level, the theoretical maximum relative infant dose expressed as a percentage of the weight-adjusted maternal dose was 0.012%. As most glaucoma patients administer drops to both eyes, the dosage was duplicated to reflect the more pertinent calculated theoretical relative infant dose of 0.024%. This dose of timolol is unlikely to cause systemic side effects to the healthy breastfed infant.