The invention provides a microfluidic detection chip, a kit and a whole blood multi-index detection method based on the same and an application of the microfluidic detection chip. The detection chip comprises two layers, the upper layer is a sample processing layer and the lower layer is a reaction layer, two or more fluid channels are arranged on the surface, facing the reaction layer, of the sample processing layer, and the two layers are closely overlapped to form a closed fluid channel system inside the chip. A plurality of biochemical indexes based on colorimetric measurement, such as liver function, kidney function, blood fat, electrolyte and the like are detected simultaneously in a small test card with a microfluidic chip technology. Difficulties in the application of the existingmicrofluidic technology are overcome, and the chip has low requirements for detection equipment, can be easily applied on site for POCT (point-of-care testing), and is applicable to large, medium andsmall medical institutions, emergency departments, community hospitals, family detection, scientific research and the like.