The invention relates to a photo-response micro-fluid self-driven micro-fluidic chip and a preparation method thereof. The surface of the micro-fluidic chip is provided with a micro-structure and a micro-channel, the surface of the micro-channel is modified through photo-response molecules, reversible changes of the surface appearance of the micro-channel and chemical configurations cause surface wettability to change under visible light stimulation, and micro-fluids are controlled to move on the surface of the micro-channel from the bottom up, so that spontaneous flow of the micro-fluids is achieved, and the micro-fluidic chip is mainly applied to related fields of electrophoretic separation, chromatographic separation, immunoassays, reactions and the like. The micro-fluidic chip is simple to operate, self-driving of the micro-fluids from the bottom up is achieved, the cost of micro-fluid driving is greatly reduced, the micro-fluidic chip has the advantages of being portable, economical, rapid and efficient, and a novel micro-fluid flow control technology is provided for non-driving flow of micro-fluids.