The mechanical properties of endothelial cells were measured using the micropipette technique. The cells employed were collected from bovine aortic endothelium and cultured in our laboratory. Endothelial cells from confluent monolayers under noflow conditions were detached from their substrate by trypsin or by a mechanical method and suspended in modified Dulbecco medium (MDM). In the micropipette technique, a part of the cell is aspirated into the tip of the micropipette under a microscope, and the deformation measured from a photograph. In this study, the data obtained were analyzed using a model where the cytoskeletal elements, which are considered to be the primary stress bearing components, are assumed to reside in a submembranous, cortical layer. Detached cells were found to have almost homogeneous mechanical properties based on measurements from different regions of the surface of a single cell. However, a hysteresis loop was observed in the relation between pressure and cell deformation during the loading and unloading processes. The calculated elastic shear moduli obtained for the trypsin-detached cells were as much as 10–20 times larger than those of a red blood cell. Mechanically-detached cells had moduli approximately twice that of the trypsin detached cells. Passage time, i.e., cell culture age, had no influence on the mechanical properties of the trypsin-detached cells, but did have an effect on the mechanically-detached cells, with both the younger and older cells being somewhat stiffer.