A micro fluidic device comprises one microstructure layer (5) and one cover layer (1), wherein the cover layer (1) is connected to the microstructure layer (5). The microstructure layer (5) comprises one bottom layer and a plurality of microstructures on it to position samples. The cover layer (1) comprises one top layer, one positioning well (6) and at least one inlet pool (4). The positioning well (6) is right above the microstructures and connected with each other. The inlet pools (4) and the positioning well (6) are connected by microchannels (3) which are formed between the microstructure layer (5) and the cover layer (1). The micro fluidic device can be applied in vitro fertilization, in determining how glial cells affect neurons, in constructing neural network and in detecting cell growth conditions.