A novel biomechanical testing methodology was developed to obtain the intrinsic material properties of an individual cell attached to a rigid substrate. With use of a newly designed cell-indentation apparatus (cytoindenter), displacement-controlled indentation tests were conducted on the surface of individual MG63 cells and the corresponding surface reaction force of each cell was measured. The cells were modeled with a linear elasticity solution of half-space indentation and the linear biphasic theory on the assumption that the viscoelastic behavior of each cell was due to the interaction between the solid cytoskeletal matrix and the cytoplasmic fluid. To obtain the intrinsic material properties (aggregate modulus, Poisson's ratio, and permeability), the data for experimental surface reaction force and deformation were curve-fitted with use of solutions predicted with a linear biphasic finite element code in conjunction with optimization routines. The MG63 osteoblast-like cells had a compressive aggregate modulus of 2.05 ± 0.89 kPa, which is two to three orders of magnitude smaller than that of articular cartilage, six to seven orders smaller than that of compact bone, and quite similar to that of leukocytes. The peremeability was 1.18 ± 0.65 (×10^-10) m⁴/N-s, which is four to six orders of magnitude larger than that of cartilage. The Poisson's ratio was 0.37 ± 0.03. The intrinsic material properties of the individual cell in this study can be useful in precisely quantifying mechanical stimuli acting on cells. This information is also needed for theories attempting to establish mechanotransductional relationships.