Cholesterol has been commonly used as a stabilizing agent within liposome formulations. However, cholesterol can have a negative impact in patients with cardiovascular diseases. Therefore, there is a need for a better stabilizing agent that can achieve liposome stability and has minimal effect on patient health. Bipolar tetraether lipids isolated from the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius, such aspolar lipid fraction E (PLFE), have been shown to make very stable liposomes. Shear rates within the body range from 40 s−1 to 2000 s−1 during physiological conditions. This study examined the effect of shear rates on dye leakage on 100 mol% DPPC, 100 mol% PLFE, 70% DPPC/30% PLFE and 70% DPPC/30% cholesterol liposomes. Liposomes with entrapped 5(6)-carboxyfluorescein (5(6)-CF) exhibit spontaneous release of the dye. Changes in fluorescence intensity were measured to monitor the leakage of 5(6)-CF. Liposomes were subjected to a constant shear rate for 3 hours through an IBIDI perfusion and pump system. Dye leakage rate constants of 5(6) CF from liposomes were determined under 0, 1000 s−1, and 7600 s−1 shear flow rates at 37°C. The data demonstrate that addition of PLFE in liposomes greatly reduces 5(6)-CF release and minimizes the effect of shear stress when compared to cholesterol. This study demonstrates that the 70% DPPC/30% PLFE liposomes were the most stable under shear stress, with 100% DPPC exhibiting a 4-fold decrease in leakage rate constant and 100% PLFE exhibiting a 2-fold decrease. Inserting PLFE into liposomal systems in place of cholesterol increases the stability of liposomes under various rates of shear stress. PLFE can potentially be utilized to improve the broad range of liposome formulations and improve the lifetime of drug release. (NSF DMR1105277 and CBET1350841, NIH/NINDS NS086570-01, The Shriners Hospitals for Children 85110-PHI-14 and NIH-NIDA T32 DA007237).