Topical, to the conjunctiva, a thin strip (approximately 0.5 cm) of a 0.025% ointment once every three days to three times a day.

Recombinant AChE was quantitated using the extinction coefficient 119, 650 M-1 cm-1 calculated from its amino acid sequence. AChE (34 nM) was assayed using Ellman’s reagent using 160 uM acetylthiocholine and 300 uM 5,50-dithiobis-2-ntirobenzoic acid (DTNB). Thiocholine released by the enzyme reacts with excess DTNB, resulting in the formation of the thionitrobenzoate anion, which absorbs at 412 nm. To assess IC50, the inhibitors were solubilized in isopropanol and used in volumes of less than 5% of the total reaction volume. AChE (34 nM) was incubated for 30 min with each compound at an array of concentrations. The residual activity of the enzyme was assayed using 500 uM acetylthiocholine, a saturating concentration 10-fold higher than the Km of 46 uM for this substrate.