The invention provides a three-dimensional display, an alignment method and an alignment system thereof. The alignment method comprises the following steps of: fixing the position of a liquid crystal grating as an alignment standard, carrying out primary alignment by adjusting the relative position of the display to the liquid crystal grating, determining the relative position between a pixel substrate of the display and the liquid crystal grating; then continuously adjusting the relative position of the display to the liquid crystal grating and carrying out more-accurate secondary alignment. Compared with a method of only matching the alignment identifier of the liquid crystal grating with the lignment identifier of the display in the prior art, the alignment method provided by the invention has the advantages that not only does an alignment identifier not need to be arranged on the display, but also the production cost is reduced; the essence of the alignment method provided by the invention is as follows: the liquid crystal grating and the pixel substrate of the display are aligned, so that the alignment accuracy of the liquid crystal grating and the display is higher and the display effect is optimal.