In recent years, the development of microfluidic technology in the fields of biology, chemistry, materials science, medicine and other fields has been particularly rapid. It has gradually shown a trend to replace the traditional methods of various functions realized in the above fields, involving cell sorting, reaction , mixing , cell location and culture  and many other multi-level applications. In particular, a variety of new coronary pneumonia detection methods and vaccine development technologies developed by domestic and foreign researchers based on microfluidic technology have played a key role in the fight against COVID-19 this year[ 6].
With the rapid development of microfluidic technology, it has gradually begun to replace the functions of traditional conventional biochemical medical laboratories and has broad application prospects [7–10].Among the many applications of microfluidic technology, the sorting function is one of the most core functions. Efficient sorting is of great significance to the fields of biology, chemistry, diagnostics, therapeutics, materials synthesis and drug development [11, 12]. The sorting method using microfluidics can be divided into active method and passive method. The passive method uses the force exerted by the particles when they flow in the microchannel to realize the sorting without applying external force to the particles. There are mainly inertial force sorting , hydrodynamic sorting , micro-structure filtering and screening  and other technologies. The other is an active sorting method, which is to apply an external force field to make different suspended particles in the micro flow channel move along different trajectories to achieve the purpose of sorting, mainly including dielectrophoresis sorting , sound force sorting [17, 18], fluorescence excitation sorting  and magnetic sorting , etc. These methods other than acoustic sorting are limited in practical applications due to their damage to biological particles or restrictions on specific conditions. When using the ultrasonic standing wave method for particle manipulation, only the particles and the medium are required to have a difference in their density or compressibility properties. The realization conditions are relatively easy, and the separation efficiency is extremely high, thus obtaining a wider range of applications.
The cells or particles in the fluid medium will be exposed to the acoustic radiation force when exposed to the sound field , and the sound force depends on the size of the cells or particles and its acoustic contrast factor (Φ). The differences in the physical properties of various cells lead to differences in the size and sign of their acoustic contrast factors, so that the acoustic radiation force can be used to separate and classify cells according to their mechanical properties. When there is an acoustic standing wave, cells or particles with positive and negative acoustic contrast factors will move to the pressure node and pressure antinode, respectively. The use of acoustophoresis to separate particles with opposite contrast factors has been widely studied. Filip Petersson et al.  successfully realized the continuous separation of lipid particles and red blood cells using standing wave ultrasound and microchannel laminar flow. They used a microchannel with a width of 350 µm and a depth of 125 µm, and it operated in a half-wavelength standing wave mode with a pressure node in the center of the channel. Since the acoustic contrast factor of the red blood cells is negative and the lipid particles are positive, the red blood cells are collected in the middle of the channel, and finally flow out from the middle channel opening and the fat particles are collected to the position of the channel wall, and finally flow out from the two side channel openings. Carl Grenvall et al.  improved the separation device to achieve the separation of somatic cells and fat particles in milk. They added two target flow channels at the entrance of the channel, and avoided the problem of particle adhesion and blockage by changing the number of nodes in the channel.
However, most cells show a positive acoustic contrast factor in an aqueous solution. In the standing wave sound field, these cells will move to the pressure node. Therefore, the use of cell diameter or the size of the acoustic contrast factor to perform cell separation methods with the same acoustic contrast factor sign has attracted widespread attention. Petersson et al.  proposed a microfluidic device for sorting based on cell size, which achieved the sorting of red blood cells, platelets and white blood cells by using acoustic methods. There is a pressure node in the center of the microchannel. They use the target flow to focus the cell flow on both sides of the channel. The cells begin to migrate to the pressure node due to the acoustic radiation force. Because the three kinds of cells receive different acoustic radiation forces, In the same time, the distance to the node is different, and the cells that experience different acoustic radiation force are separated and collected at different positions of the exit. Tony Jun Huang et al.  designed a new structure to sort white blood cells. They tilted two pairs of interdigital transducers with the flow channel at a certain angle, so that the particles have a greater migration distance, which solves the problem that the migration distance of ordinary structures is limited by the distance between pressure nodes when sorting. Augustsson et al.  used two acoustic standing waves to separate prostate cancer cells and white blood cells. They used two sets of standing waves at the upstream and downstream of the microchannel, the first set of standing waves focused the cells on both sides of the flow channel wall, and the second set of standing waves made the cells migrate to the central node. Two types of cells with different final volumes are collected at different outlets. Harris et al.  proposed using three acoustic standing waves with frequency switches to separate cells. Using the different speeds of the cells to pressure nodes at different positions at different frequencies, by switching the different acoustic field resonance modes of the channel to separate cells of different sizes or contrast factors, this design greatly reduces the restriction on the residence time of the target cells in ordinary devices .
In this article, we propose a two-stage particle separation channel based on standing wave surface acoustic waves (SSAW) to separate three kinds of particles with positive acoustic contrast factors in microfluidic channels. The fluid dynamics focusing technology is used to control the initial position of the particles, and the separation of the particles is achieved through two different sound pressure regions. The first section is the low pressure zone, which is used to separate 9.4µm particles, and the second section is the high pressure zone, which is used to separate the remaining two particles with a smaller volume difference.
This paper studies in detail the process of particle separation using acoustic surface standing wave devices. First, we selected piezoelectric substrate materials, and compared the properties of three common piezoelectric materials qualitatively and quantitatively. Subsequently, the influence of the number of electrode pairs of the interdigital transducer and the electrode voltage on the output acoustic wave was studied. After selecting the corresponding appropriate parameters, the corresponding separation model was constructed and simulated, and the separation of three different sizes of particles was successfully achieved.