The invention belongs to the technical field of a biological chip, and discloses a micro-molecular chip, its structuring method, an application and a detecting method. The micro-molecular chip is formed by chemically bonding a light crosslinking chip and a micro-molecular compound, wherein the micro-molecular compound can be combined with Y frame protein 18 (SOX18 protein) in a sex determination zone; the invention further provides a structuring method of the micro-molecular chip; the micro-molecular compound by the method is high in fixing efficiency and good in fixing effect; the invention further provides a method for detecting SOX 18 proteins by the micro-molecular chip; the method is simple in operation and accurate in detection result. The micro-molecular chip provided by the invention is used for detecting the SOX18 protein in the sample, and has advantages of low cost and stable performance.