This study reports the use of electrorotation (ER) technique within a micro-fabricated device with 3D electrodes for dielectric characterization of Imatinib-resistant K562 (K562/IMA-0.2) human leukemic cells. The ER devices with 3D quadruple electrodes (30 mu m in height) were used in order to eliminate the fringing field effect on the rotation of cells. To induce the rotational moment on the cells, signals in phase quadrature were applied to the triangular electrodes. The rotation of cells measured at the frequency range of 1 kHz to 0.1 MHz in various medium conductivities. The membrane effective conductance, membrane capacitance, and the inner conductivity of cells were measured as 2953 +/- 82 [Sm-2], 15.63 +/- 3.02 [mFm(-2)], and 0.60 +/- 0.10 [Sm-1], respectively. These results can be used in the designing and implementation of MEMS based DEP devices for separation and detection of imatinib resistant K562 cells.