The fluorescence polarization of A23187 is used to detect physical changes in myoblast membranes produced by Ca2+ concentrations which are able to trigger fusion. Temperature scans reveal a Ca2+-dependent fluidity increase in the microenvironment of the ionophore above 37 degree C. 2. Time-dependent polarization measurements show two different effects. The first one consists of a fast polarization increase which reaches its maximum after 5-10 min. This change could be explained by a Ca2+-induced phase separation of acidic and neutral phospholipids in the membrane. The second effect is a slow polarization decrease over 2-3 h subsequent to the fast increase. 3. The information derived from the fluorescence polarization is in accordance with the characteristics of the fusion of myoblast cells regarding the dependence on Ca2+ concentration, the inhibition by Mg2+ and lysolecithin as well as a typical temperature of 35-37 degree C where fusion rate changes abruptly.