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In this paper, laser interference induced dielectrophoresis (LIIDEP) was presented for cell manipulation and three-dimensional array assembly. Two-beam and three-beam interference systems were built to manipulate yeast cells in deionized water. When the laser power was 60 μW and the sinusoidal AC signal was 6 V at 10 kHz, the two-beam interference pattern manipulated the cells into a single-row array distribution. For three-beam interference, when the laser power was 70 μW and the frequency was 4.5 kHz, the voltage of 2.5 V enabled each spot to control one cell, and 7.5 V enabled many spots to control multiple cells. When the cell concentration was 2 × 107 cells/mL and the electrical signal was 8 V and 4.5 kHz, the cells were assembled into a three-dimensional columnar array. The feature size of laser interference pattern was adjusted to microscale and maintained sharp boundaries, which effectively improved the manipulation accuracy. Compared to the projector or laser as the ODEP light source, LIIDEP has the advantages of high manipulation resolution, good contrast and large dielectrophoretic force. This work provides an effective way for high resolution three-dimensional array manipulation and assembly of a large number of living cells on the microscale in liquid environments. • The interference pattern effectively improves the precision of cells operation. • Two-beam interference pattern manipulated cells into a single-row array distribution. • Each spot of the three-beam interference pattern controls single or multiple cells. • The cells can be quickly assembled into a 3D columnar array. [ABSTRACT FROM AUTHOR]