A three-dimensional stereoscopic imaging method includes: longitudinally slicing a three-dimensional scene according to a physical depth of field to produce several two-dimensional layers; respectively displaying each two-dimensional layer on each different two-dimensional display window to form several two-dimensional pictures; polarizing light emitted from the two-dimensional display windows to transmit the light into human eyes, in such a manner that the human eyes are able to perceive the images of all the pictures, and splicing the images of all the two-dimensional pictures into an image of the original three-dimensional scene. The method realizes a three-dimensional stereoscopic imaging by simulating a real physical depth of field generated by human eyes perceiving a real three-dimensional stereoscopic image. Viewers are able to see a three-dimensional scene without wearing glasses. A three-dimensional stereoscopic imaging system and an imaging device thereof are further provided.