Kinetics of calcium phosphate-induced fusion of human erythrocyte ghosts monitored by mixing of aqueous contents.

We have adapted the terbium fusion assay (Wilschut, J. and Papahadjopoulos, D. (1979) Nature 281, 690-692), which has proven to monitor the mixing of internal contents during phospholipid vesicle fusion in a reliable manner (Hoekstra, D. (1982) Biochim. Biophys. Acta 692, 171-175), to study the fusion of erythrocyte ghosts as induced by the combined action of Ca2+ and phosphate. Using this assay, it became possible to reveal, for the first time, the kinetics of fusion of a biological membrane vesicle system. The rate of fusion was critically dependent on the concentration of Ca2+ and phosphate. Prior addition of phosphate was essential for induction of fusion. Initial fusion was largely non-leaky, but in a process secondary to the fusion event the ghosts gradually released their contents. It is suggested that the experimental approach presented in this paper, would facilitate efforts to elucidate the mechanism of fusion of biological membranes.