In this paper, citric acid was used as carbon source for preparing fluorescent carbon quantum dots(CQDs). Due to the obtained CQDs had carboxyl groups on their surfaces,Man-CQDs were synthesized by immobilizing 4-aminopheny-l-α-D-mannopyanoside on the surfaces of CQDs via covalent bond.Furthermore,the binding constants of Man-CQDs and Escherichia coli JM109/Escherichia DH5a/Salmonella typhimurium S.123443 were obtained by fluorescent competition experiments.The results showed that the sizes of CQDs were approximately 26nm,the maximum excitation wavelength for CQDs was 445nm and the fluorescence yield was 76% in contrast to 54% quinine sulfate. Compared with the fluorescence intensity of CQDs,the prepared Man-CQDs remained unchanged in principle. By the Phenol-sulfate method, the concentration of Man-CQDs was calculated to be 2.832mmol/L,and the content of mannose in Man-CQDs nanoparticles was approximately 40%.Based on competition binding experiments of Man-CQDs and D-mannose with bacteria, the binding constant of Man-CQDs and bacteria was calculated by Scatchard equation, namely the binding constants of Man-CQDs with Escherichia coli JM109 and Salmonella typhimurium S.123443 were 2.39×103L/mol and 1.17×105L/mol,respectively. However, the reaction of Man-CQDs and Escherichia coli DH5a was nonspecific binding. The results suggested that the proposed method was applicable for the determination of noncovalent binding constant between mannose and bacteria. Besides, it offered a valuable reference information for the research of interaction between carbohydrate and bacteria.