The invention discloses a multi-channel micro-ring sensor and a preparation and use method thereof, the multi-channel micro-ring sensor comprises a sensing substrate, a waveguide micro-ring, a coating exposed sensing area and an optical fiber array, the waveguide micro-ring is manufactured on the sensing substrate by adopting a standard tape-out process, and the waveguide micro-ring comprises an input waveguide, two output waveguides and a micro-ring; according to the present invention, the multi-parameter detection purpose which can only be achieved through the traditional micro-fluidic channel is improved to the combination of the parylene C film hydrophobic effect and the microspotting instrument, such that the specific protein is fixed on the micro-ring surface through the one-step method so as to simplify the chip preparation and modification process, and the traditional micro-fluidic channel is adopted such that the size of the chip cannot be too small. The method does not depend on a microfluidic channel, the size of the chip is reduced, the higher integration level is achieved, meanwhile, the parylene C thin film one-step method is used for fixing the specific antibody, after detection is completed, parylene C can be removed, the chip is recycled, and the cost is reduced.