The invention discloses a fluorescent probe for detecting and quantifying signal molecule of Gram negative bacteria colony, and belongs to the technical field of rapid detection of pathogenic bacterium. According to the fluorescent probe, a quantum dot is connected with MIP through a surface molecular imprinting technology, and the purpose of accurate quantitative detection is achieved through a fluorescence value on the basis of high degree specificity detection and fluorescence. MIP polymer is precipitated and polymerized on the surface of an amino-silane functional group modified by a CdSe/ZnS quantum dot in advance, so that QDs@MIP is successfully synthesized. The invention further discloses a preparation method and the application of the molecular imprinting fluorescent probe. The fluorescent probe can realize indirect detection of part of Gram negative bacteria, has the advantages of sensitivity, speediness and high specificity, and is low in price, and the method has an important implication for food safety or health care facilities, and can be applied to the direct monitoring of Gram negative bacteria.