Using micropipette aspiration and fluorescence techniques, we have studied the material properties of charged lipid vesicles in calcium solutions. Vesicles were composed of phosphatidylglycerol (PG)/phosphatidylcholine (PC) or phosphatidic acid (PA)/PC mixtures. For the case of PG/PC membranes, we measure no effect of anionic lipid fraction on elasticity but a monotonic decrease up to 20% for tension required to induce membrane failure. Both of these observations are rationalized by a model we have developed to describe membrane electrostatic interactions in a two-component salt solution and the resulting changes in membrane properties. Critical tensions measured for PA/PC membranes, on the other hand, did not depend on anionic lipid fraction and were uniformly similar to35% lower than PG/PC vesicles. This is likely due to a lateral phase separation in the membrane. By combining mechanical properties with fluorescence observations we propose that the PA-rich phase separates into small unconnected domains. (C) 2003 Elsevier Inc. All rights reserved.