Liposomes containing acetylcholinesterase were prepared by the freeze-drying method. The multilamellar morphology of the vesicles was revealed by freeze-fracture electron microscopy and their size distribution was determined by quasi-elastic light scattering. The vesicle diameters were in the range of about 0.2-4.0 micron. The liposome preparations were tested for their ocular delivery of an entrapped cholinesterase enzyme in counteracting the miotic effect of diisopropylfluorophosphate (DFP), a prototype of a family of organophosphate poisons. The topical application of the enzyme-containing liposomes to the rabbit eye was found to confer a significant level of protection against DFP-induced miosis. In comparing the prophylactic effectiveness of different enzyme-bearing liposomes, positively charged vesicles were found to be more effective than either neutral or negatively charged vesicles. Although the precise protective mechanism is not clear, our in vitro studies indicate that DFP molecules freely associate with liposomes and tear fluid promotes the release of liposome-entrapped enzymes. Thus, it is conceivable that the enzyme-liposome complex may act somewhat like a sponge by sequestering DFP molecules which diffuse into the vesicle, and also by releasing the entrapped enzyme to combine with DFP, thereby neutralizing its in vivo toxic effect.