We present a measurement of the electric charge of top quarks using tt¯ events produced in pp¯ collisions at the Tevatron. The analysis is based on fully reconstructed tt¯ pairs in lepton+jets final states. Using data corresponding to 5.3  fb-1 of integrated luminosity, we exclude the hypothesis that the top quark has a charge of Q=-4/3e at a significance greater than 5 standard deviations. We also place an upper limit of 0.46 on the fraction of such quarks that can be present in an admixture with the standard model top quarks (Q=+2/3e) at a 95% confidence level.