Recently, microfluidic Devices have been used for the separation of particles and cells based on their capability such as high efficiency[Adams, 2012 #147], the ability to analyze small sample of particles and potential for clinical treatments. Precise manipulation of particles and cells within microfluidic channels is usually based on dielectrophoresis, inertial focusing, magnetic tweezers, or acoustophoresis. In the present study, we investigate and simulate the combination of two separator methods in a spiral microchannel (a kind of inertial separators) and separator in the microchannel under the influence of acoustic wave in three dimension, using comsol software. In fact, the spiral microchannel sectio is a pre-separator for microchannel under the influence of acoustic wave. In the spiral microchannel section, an initial separation is performed, then the particles enter the microchannel under the influence of acoustic wave and the final separation of the particles is done in this section. The governing equations of acoustic waves are obtained by creating a small perturbation in the steady state Navier stokes equatio therefore, one simple way of modeling acoustic waves is to use first-order perturbation theory. On the other hand, spiral microchannels also cause particle to attain new equilibrium position ased on their geometric and mechanical properties, by creating lift and drag forces, Which is the cause of the spiral geometry. The purpose of this study is to investigate the separation of particles with 10 , 20 and 30 diameter and constant properties from each other in a fluid, and its use for separation of particles and biological cells through acoustic and hydrodynamic forces in the blood. This simulation is in three dimensions and is done for tow methods(solving governing equations of acoustic waves by Gorkov force method and solving Helmholtz equations by Gorkov force method) for acoustic waves. In this study, the effect of particle collisions with each other and the impact of particles on the walls of the microchannel is not considered. In addition, it is assumed that the motion and forces applied to the particles do not affect the fluid flow and the acoustic field. Many parameters affect this separation, including the Dean number, the frequency of the acoustic wave, and the diameter of the particles. Due to the properties of the particles and the chose dean number, increasing the spiral microchannel loops, the particles are transmitted to the upper part of the central axis of the microcanal. On the other hand, these particles move under the influence of acoustic force toward the acoustic pressure nodes. Therefore, by changing the number of spiral microchannel loops, some particles are separated from the other particles by the spiral microchannel section. Then, when the particles arrive into the part under the influence of the acoustic wave, they are separated further by the acoustic force. The results show that the separation of particles from this hybrid system can also be accomplished with a 100% separation rate. Keywords: Acoustic wave, Spiral microchannel, Micro particles, Constant Properties