We have devised a simple method of determining the time course of the effects of timolol maleate on aqueous flow using changes in aqueous protein concentrations. Aqueous protein concentrations were measured with a laser flare-cell meter. First, the value of the coefficient of protein entry into the anterior chamber (kin) was determined in both eyes of 10 normal subjects. One week later, aqueous protein concentration changes were measured in both eyes of the same subjects before and after instillation of timolol in one eye. From these changes and the value of kin, changes in the aqueous flow rate were calculated using an equation formulating protein kinetics in the anterior chamber. Results of a separate experiment showed that instillation of topical timolol did not change the kin, justifying use of the kin as determined without considering timolol in the calculation. The calculated flow rate had a maximum decrease of 30% +/- 10% (mean +/- SEM, n = 10) 4 hours after instillation in treated eyes and of 10% +/- 6% 3 hours after instillation in fellow, untreated eyes.