The mechanical characteristics of non-ionic bilayer membranes composed of sorbitan monostearate, cholesterol and poly-24-oxyethylene cholesteryl were studied by measuring the modulus of surface elasticity ( μ), a measure of membrane strength, as a function of cholesterol content and temperature. The modulus of surface elasticity increased slowly with increasing cholesterol concentration, with a sharp increase around 40 mol% cholesterol (on average an increment of 0.43 × 10 6 Nm −2 per molar percentage), and displayed a maximum of 6.5 × 10 6 Nm −2 around 47.5 mol% cholesterol. Further cholesterol resulted in a decrease in μ. Generally the interaction of cholesterol with the sorbitan monostearate should increase the rigidity of the membrane. However, the latter effect may be due to the formation of cholesterol clusters at high cholesterol content where excess amounts of cholesterol cannot interact with the sorbitan monostearate, and deposits on the bilayers compromising their uniformity, strength and permeability. This behaviour was evident when measurements were carried out above and below 25 °C.